What are the problems in using an ssDNA or an mRNA for expression instead of using a plasmid?

What is the problem with handling pure mRNA nucleotides of gene X? Or
  even sscDNA?

Problems in directly transfecting a mRNA/ssDNA

mRNAs can be transfected into the cells and is quite frequently done. The problem with transfecting mRNAs directly is that the mRNA gets degraded soon and you will not have sustained expression. This is, however, very useful in certain cases such as creating induced pleuripotent stem cells (iPSCs) where you do not want the cells to continually express the Yamanaka factors (some of which are also oncogenes).

Also, RNAs as such are quite susceptible to degradation. It is much easier to prepare and store DNA solutions for a long time.

Similar problems arise with a ssDNA.

  Why is it you can only buy X already inside a vector (which is in turn often inside a bacterial carrier)?

One great advantage with using plasmids is that you can propagate your gene infinitely. If you were instead given a fixed amount of mRNA, then you will run out of it soon. Some vendors do practice such greedy capitalistic policies :P

When you are amplifying a DNA using PCR then cloning it has an additional advantages:

you can sequence the gene using well standardized primers (like T7)
you can make constructs that can express the gene conditionally

  You have an empty expression vector (mRNA or DNA) and are looking to
  clone some gene X into it.

You cannot easily clone something inside an RNA. You need specific RNA endonucleases. Moreover a double digestion will be disastrous in this case.

  Why vendors don't generally sell a dsDNA insert but rather provide it cloned in a plasmid

A circular dsDNA is more stable
Vendors can keep the stock of the insert for future customers without having to do PCR again
Cloned insert can be amplified via bacterial cultures to obtain very high amounts (maxi/giga preps)


Fish ID? Small, yellow-finned, tropical fish in Thailand

With a little Google search I found similar fishes. Says it is golden or yellow damselfish or possibly lemon damsel but I'm not sure because there are some different names for this picture or different pictures for searching same name can be found on Google.

Also, I found a fish identification page on which this fish can be found.

Is this your fish?


Explanation of ECG in MI using an electrostatic model

The concentration of K+ inside the cell is 30 time more than outside. Hypoxia causes loss of this intracellular Kalium and consequent/premature repolarization when measuring the potential of action.

As the all the ECG theory is Vector based, any abnormality in the electric pathways and myocytes will result in changes of ECG.

This picture depicts the ST changes due to different extension of injury. In addition, the explanation on this site will probably resolve the issue of understanding.

ADD

The article explains the ECG phenomena as a result of electric propagation of dipoles. If the dipoles propagate towards the lead we get upward wave etc.

As the electrostatic model takes the whole heart as one dipole (as a sum of multiple dipoles) the minor infarcts can be missed on ECG. It is an actual fact, this is why other tests are important (for example, the troponin test).

The confusion point in your understanding is a "dead" tissue. In a case of infarct there is an hypoxic damage which leads to elevated permeability of the cell membrane with resulting K leak (as I explained above there are 30 times more Kalium inside the cell than outside). This means that "dead" area will have abnormal depolarization/repolarization pattern just because there are multiple "non grouped" dipoles in that area (look at the picts). There are diastolic and systolic theories of ST elevation, but none of them has won the battle yet.

With time the scarring tissue replaces the infarcted myocardium and in that case the ST-segment will not be affected and will look nearly normal. This is the case you meant as "dead" tissue - scar tissue does not leak any electrolytes and consequently the ST segment is not affected in a chronic stage.


Are all body atoms really recycled several times during a life?

Rudolf Shoenheimer and David Rittenberg were key figures in introducing the isotopic tracer technique to biology.
This technique, which may be dated to the discovery of deuterium by Urey in 1933, produced a revolution in biological thought.
Much of the early work was done at the Department of Biological Chemistry at Columbia University under the chairmanship of H. T. Clark.
Up to this time it was thought that the components of tissues were relatively stable, and that ingested food was metabolized immediately to provide a source of energy.
The first experiments showed that when deuterated fatty acids were fed to mice, most of the label was initially deposited in adipose tissue.
It was concluded that adipose tissue acts a 'fuel depot' that is in a state of metabolic flux and is the direct source of fatty acids burned as a fuel (see here for refs). Fats present in food are not primarily used an an immediate energy source.
When a stable isotope of nitrogen became available (N15), it was found that proteins were also 'in a state of dynamic flux' and were being continuously regenerated and broken down.
The first experiments were done with N15-labeled tyrosine and it it worth quoting from the paper (Studies in Protein Metabolism. VII The metabolism of tyrosine by D Rittenberg Rudolf Schoenheimer &  S. Ratner)

The results indicate that in our rat the nitrogen of at least one
amino acid, tyrosine, was only partly excreted in the urine, while
almost half of it was retained in the body proteins.
Of this deposited nitrogen only a fraction was found attached to the original
carbon chain, namely to tyrosine, while the bulk was distributed over other
nitrogenous groups of the proteins

Shortly after giving a series of lectures at Harvard University in 1941, Shoenheimer took his own life.  The lectures were published posthumously as a classic book called The Dynamic State of Body Constituents.
Further information about the early  work on isotopes, their role in changing completely the way we think about metabolism, and the role of Columbia University in shepherding in this great era, may be found in the following open-access article by Eugene. P. Kennedy:
Hitler’s Gift and the Era of Biosynthesis
Another good reference on the work of Schoenheimer may be found here

So are all proteins broken down and regenerated?  I know of any one exception and that is the lens crystallins where there is virtually no turnover
throughout the life of an individual.
The crystallins in our lens are synthesized at birth and remain with us virtually unchanged for life.
There is a very nice open-access paper on this published in PLOS One:
Radiocarbon Dating of the Human Eye Lens Crystallines Reveal Proteins without Carbon Turnover throughout Life
(Lynnerup et al. 2007)


Tree Identification - California Bay Area

For California trees, my go-to is A Californian's Guide to the Trees Among Us by Matt Ritter (Heyday, 2011).  California trees can be challenging due to the extremely varied habitat and many introductions over the years.
Your trees appear to be younger specimens of Pinus pinea or the Italian Stone Pine, based on the description on page 14 in Ritter. No other conifer in the book matches better.  Going to Wikipedia the description also seems to fit, in particular they mention "...in youth it is a bushy globe..."  There is also good information at Kew where I found this image, which looks to be very similar to what you observed.
Of course, if you can visit the location again you can check for needles in pairs, and orange-brown scaly bark.


What are some uses of oxidative/reductive power inside cells?

This topic is quite broad, excluding energetic metabolism, there are still many reactions that use redox equivalents, whether NADH, NADPH, or FADH. Here are some paradigmatic illustrations.

Xenobiotic metabolism

Oxydation of substrate : as you mentioned, alcohol to aldehyde to carboxylic acid. Generate reducing power. (The alkane oxydation shown below occurs in bacteria)

Mono-oxygenation : hydroxylation of a substrate with $O_2$, with the other oxygen atom reduced to water. This proceeds as follow with CYP450 enzymes :

Regeneration of oxided gluthatione

Anabolic metabolism

Basically every time there is a need to create/remove a double bond, or move from carbonyl to aldehyde to alcohol and back, redox equivalent are used.

Fatty acid elongation : as this proceeds with acetyl-CoA as carbon source, intuitively one sees the need to reduce the carbonyl to obtain an alkane chain.

HMG-CoA Reductase (HMGCoAR) catalyses the commited step in cholesterol synthesis.

Synthesis of prostaglandins from arachidonic acid. (More generally, any lipidic mediator)

There are many, many more examples. You may want to check out the Wikipedia page for NADH, NADPH, FADH, or the list of enzyme that use them (1, 2, 3)

All illustrations from Biochemical Pathways, G. Michal & D. Schomburg, 2nd edition, Wiley


Disclaimer

As I have pointed out in my comment, it is not clear whether the sources mentioned relate to eukaryotes or prokaryotes, assuming they are correct. I am a translation man, rather than a transcription man, and so am answering this from the 2002 edition of Berg et al. ‘Biochemistry’, as I happen to have a copy of my own (a freebie from when I was still teaching) and this edition is freely available online. List members with more expertise in this area are encouraged to post comments to correct any errors in my answer or supply updates.

Answer

in neither prokaryotes nor eukaryotes is the DNA helicase that operates during DNA replication involved in transcription although other proteins necessary for transcription have DNA helicase activity.
In prokaryotes it appears that the RNA polymerase holoenzyme (made up of just four subunits) is responsible for unwinding about 17 base-pairs of template DNA. Quoting from Section 28.1.3:

  Each bound polymerase molecule unwinds a 17-bp segment of DNA, which corresponds to 1.6 turns of B-DNA helix

There may be some ambiguity caused by the description of how this value was determined experimentally as it involved addition of topoisomerase II. However it is clear from discussion elsewhere that this is an enzyme of DNA replication and is not involved in transcription.
In eukaryotes transcription is much more complex, involving separate polymerases for different classes of RNA product and a variety of auxiliary transcription factors in the case of RNA polymerase II, which transcribes mRNA. According to Section 28.2.4:

  The TATA box of DNA binds to the concave surface of TBP [the TATA-box-binding protein]. This binding induces large conformational changes in the bound DNA. The double helix is substantially unwound to widen its minor groove, enabling it to make extensive contact with the antiparallel β strands on the concave side of TBP.

So the initial recognition of the TATA box causes some unwinding. However the major player appears to be transcription factor TFIIF:

  an ATP-dependent helicase that initially separates the DNA duplex for the polymerase


The chromosomes in your picture do have sister chromatids, they are just very close together. A chromosome needs to pass through replication before it can compact into the typical metaphase chromosome shape depicted above. When I prepare metaphase chromosomes, I usually see a mixture of chromosome sets where the chromatids are close together (like above) and clearly separated. I think this is related to the time each metaphase has spent in the colcemid block-the longer the cell has been blocked at metaphase for, the longer the enzymes that work to separate the two chromatids have acted on it. Also chromosomes keep getting more and more compacted during the colcemid block-if you leave them for too long, they become a lot smaller.  This paper has very cool pictures showing the links between the sister chromatids:
http://www.ncbi.nlm.nih.gov/pubmed/18461479


As has been commented, the answers to some of the questions here can be found in the literature. The nutrients and oxygen reach all tissues by the circulatory system — i.e. in the blood. And the link supplied by @trondhansen suggests a litre of oxygen a day. (I wouldn’t know — I never studied physiology.) However, as regards energy, I supply the following from the biochemistry text, Berg et al.

Brain. Glucose is virtually the sole fuel for the human brain, except during prolonged starvation. The brain lacks fuel stores and hence requires a continuous supply of glucose. It consumes about 120 g daily, which corresponds to an energy input of about 420 kcal (1760 kJ), accounting for some 60% of the utilization of glucose by the whole body in the resting state.

Note the correct form of expression:

the brain  / consumes fuel / to generate energy / to perform ‘work’

Glucose is the fuel, ATP is the chemical energy currency, and the work performed (as the text makes clear) is for the main (60%–70%) to:

maintain the sodium/potassium ion membrane potential required for the transmission of the nerve impulses.

(I am afraid that’s about as layman’s terms as I do. It does say on the label that this site is for students of biology.)


Nice question! It would, obviously, be a long answer if I talk about all types of sulfur compounds here. So I will take up just 2 examples to explain this.

Thiols: when we talk about sulfur compounds with foul smell, thiols (mostly) come to the top in the list. Thiols (R-SH) are a class of compounds famous for their smell. Some examples of thiols include:

compounds in onion and garlic (see this answer for more details).
a mix of compounds responsible for the smell of skunk's spray (Anderson et al, 1975)
(R)/(S)-3-methyl-3-sulfanylhexan-1-ol found in human sweat
(Methylthio)methanethiol found in male mouse urine

Some biologically important examples would include coenzyme-A, glutathione and cysteine. However, not all thiols have fowl smell. For example, furan-2-ylmethanethiol provides the aroma of roasted coffee while grapefruit mercaptan gives grapefruit its characteristic scent. You can see a complete list here.

After seeing some examples, lets come to the main point i.e. why thiols have such foul smell. Thiols are decomposition products of proteins. When amino acids, like cysteine and methionine, are decomposed, thiols are one of the last things to form. This would explain why it becomes important to smell and avoid them (you wouldn't want to eat a dead and decaying mammoth, and a blind hunter might not be able to tell if he has found a decaying mammoth if he can't smell such compounds). As a side note, this also explains why organic acids have sour taste and smell. Compounds like glucuronic acid, citric acid, oxalic acid, lactic acid, butyric acid, acetic acid, etc. are all decomposition products (if you know how vinegar is made traditionally). This is why your body tries to get rid of them (I saw that in a video, if you eat things with foul smell and sour taste (it was a Japanese dish probably), you'll suffer nausea and vomiting since your body will do all it can to prevent its harmful effects). See this Wikipedia page for more info.

Hydrogen Sulfide: H2S, or hydrogen sulfide, is another compounds famous for its smell. It smells like rotten eggs, and indeed it is because H2S is what rotten eggs release. The basic reason why H2S smells so bad is again the same. Hydrogen sulfide is one of the final products of proteolysis during decomposition (see same Wikipedia article). But there is another cause why H2S has such foul smell (and why I didn't just stop at thiols): H2S is itself toxic.

H2S has many biological signalling functions, similar to NO and CO. These include:

H2S serves as endotheium-derived relaxing factor (EDRF) and endothelium-derived hyperpolarizing factor (EDHF). In short, it acts as smooth muscle relaxant and vasodilator. It also increases the response of NMDA receptor and facilitates long term potentiation in the brain.
it is converted to sulfite by thiosulfate reductase, and further to thiosulfate and sulfate by sulfite reductase, in the mitochondria. Sulfate is excreted in the urine.
it acts on ATP-sensitive potassium channels in smooth muscles, and does the blood vessel-relaxing work in smaller blood vessels.
it blunts, reverses and promotes healing of diverse inflammatory reactions. A full list is available on this Wikipedia page.
the most pronounced effect of H2S is much similar to that of CO. It also binds to the iron in the mitochondrial cytochrome c oxidase and other mitochondrial cytochrome enzymes, thus preventing cellular respiration and causing death.

This would explain why external source of H2S would be so harmful since, being biologically such an active molecule, its amount needs to be controlled strictly. Now, if you want to know why decaying bodies would be dangerous, this Wikipedia article would be a good starting point.

EDIT: The bitter taste of some vegetables is because of a type of compounds called glucosinolates found in some plants. These compounds dissociate upon eating to form isothiocyanate and are thought to be a part of the plants' defense system (see this answer for more details). Interestingly, a similar type of compound, cucurbitacin, found in Cucurbitacae plants, does not contain sulfur, yet it is responsible for the bitter taste of these plants.


Why do we need to put phosphate fertilizers along with rhizobium?

Phosphorus is indeed a macronutrient!!! If you're wondering why it is insufficient in natural soils then read on. Phosphorus is, in fact, the most limited of the three primary macronutrients in most natural soils (being nitrogen, phosphorus and potassium). This is because it is released very slowly from insoluble phosphates (on rock surfaces) and rapidly becomes “fixed” after being released, making it difficult for plants to obtain their phosphate ions (intake form of phosphorus). Few unfertilized soils release phosphates fast enough to support the high growth rates of many plant species. Also, plants tend to only obtain their phosphate ions from the little “polyprotic phosphoric acid” in the soil, they are often very deficient in phosphorus and for this reason phosphate ions are used in high quantities than nitrogen and potassium in NPK fertilisers (N:P:K = 1:2:1).

P is found in natural compounds in soil (however this is very limited); it is more often found in NPK fertilisers in the forms of: Orthophosphates ( H2PO4- or HPO42- ) and Polyprotic phosphoric acid ( H3PO4 ).

The following reasons are why phosphorus is a primary macromolecule and why it is VERY essential to plant nutrition;

To build ATP (the energy currency of plants) from respiration ;
To build DNA and RNA (genetic information) ;
To build the cell membrane ;
To aid the growth of a plant in its early stages (seedlings and root development) ;
To perform phosphorylation ( the modification of the activity of various enzymes – this is responsible for cell signalling (which is the ability of cells to perceive and correctly respond to their microenvironments – aka tissue homoeostasis))


Interlocus Contest Evolution means suicide?

X and Y chromosomes don't "want" anything-they are not conscious. I know that sounds obnoxious, but it's very important in evolutionary biology to understand that there is no agency and no intended final product for natural selection. I think the best way to think about it is like this: let's say there's a mutation on the X chromosome that happens to make females more fit at the expense of making males less fit. It seems possible that the evolutionary arms race that this establishes (i.e., males bearing a mutation that compensates or overcompensates for the new expense of the mutation that, say, their successful mom gave them, would then start to sweep to fixation in the population, and so on and so on) could result in population collapse. Apparently, there's theory out there that confirms this: http://www.els.net/WileyCDA/ElsArticle/refId-a0003669.html. Also, I vaguely remember a paper about toxic sperm (vs. female mating behavior) in Drosophila melanogaster suggesting that populations could collapse. As Bonduriansky mentions in the link, though, one resolution to the arms race would be the evolution of genes that get expressed very differently between the sexes (i.e., a gene that is good for a female but terrible for a male would evolve to only be turned on when it's found in a female). I hope that helps?


Does loss of dopaminergic neurons totally eliminate voluntary muscular control?

Suppose that someone has completely lost the ability to voluntarily control his/her muscles due to the destruction of the brain's dopaminergic neurons in the substania nigra

This is not correct. The substantia nigra plays an important role in both reward and movement. The degeneration of dopaminergic neurons in the substantia in Parkinson’s disease leads to loss of fine motor movement (leading to limited movement and tremors) and some loss of non-motor skills (loss of smell).

What locations control breathing?

The autonomous regulation of breathing is controlled by bundles of neurons called the dorsal respiratory groups (DRG) and ventral respiratory group (VRG) that together make up the medullary respiratory center. Source.

Though the exact mechanism of how we control our voluntary breathing is still a debated. We have a general idea:

  Voluntary control of breathing is mediated primarily via the
  corticospinal and corticobulbar tracts and is important in activities
  such as speech, singing, and voluntary breath-holding. Disorders
  involving this mechanism occur mainly in bilateral pontine infarctions
  and lesions involving the pontine tegmentum that interrupt the
  descending motor pathways. - Neurology and Clinical Neuroscience, 2007

Would that person be able to breathe manually if he/she so wished, or would he/she only ever breathe autonomically?

Although the exact mechanism for voluntary breathing is not fully understood, the voluntary process is controlled by a series of nerve bundles. If the nerve bundles were severed, there would be decreased voluntary control over the breathing process. Losing the ability to manually breath could be bad for overall health. One study has found that a lack of the ability to manually breath can lead to poor spinal stabilization and posture.

What if you lost autonomous control?

There is a rare disorder known as central hypoventilation syndrome (CHS) that results in respiratory arrest during sleep (also known as Ondine's Curse). This disorder is caused by a lack of development of the neural tube (aka Medulla and the VRG/DRG) during birth or some type of traumatic brain injury. Treatment for this disorder is a lifetime of mechanical ventilation.


What does it mean by “end effects can be ignored”

They are considering a model of a DNA sequence of quasi infinite size. They consider a case where the DNA sequence is so long that the fact that it has ends will only negligibly affect the conclusions of their model. This is likely the same as considering the DNA is circular.

To give you a similar logic, imagine a species where individuals live in different populations. All population can be placed on a one dimensional axis. The migration rate from one population to the neighbouring population is 0.1 (this is called a "stepping stone model" btw). Hence, the probability of not migrating is 0.8 (1 - 0.1 - 0.1). I can draw conclusions from such a model but it would be important to specify that I neglected the end effects and hence, I assumed there are a lot of populations (or that the population form a loop). Why did I neglect the end effects. The last population on each side cannot have a migration rate of 0.1 toward a population that does not exist. Hence, their migration rate must differ from the other populations. This is commonly called edge effects (rather than "end effects")


Why is absorbance at 280 nm for protein solution going up when I measure repeatedly?

It looks like your protein concentrations are right on the limit of detection of the spectrophotometer, and changing the diluent buffer changed their concentrations. The samples may not have been thoroughly mixed after dilution and before measurement, so the varying measurements may simply be the solution coming to equilibrium. Temperature can also affect absorbance, so you should verify that your samples have equilibrated before drawing any conclusions. If the absorbance of your phosphate buffer is 0.03, I'd try to keep the sample absorbances above 0.075 or higher to avoid getting too close to the limit of detection. Also, make sure your buffer isn't too old or contaminated with something which could be affecting its absorbance characteristics.

I would suggest taking one or two protein samples and doing a dilution series (1:1, 1:5, 1:10, 1:20) in a large-ish volume (say 400 ul each, if you can spare it), vortex briefly to mix well, then measure triplicates of each dilution on your reader, along with appropriate blanks (buffer only). You will see differences between each measurement, but it should be quite small, depending on the accuracy and precision of your instrument.

Measured values will not be exactly the same from measurement to measurement, and it would take a lot more than three repetitions to determine if there was an actual drift trend occurring. Measure your sample plate every 5 minutes for an hour and plot the values (don't just eyeball them) to see if the machine may need to be serviced.


How to calculate the occurrence of a stretch of nucleotides in a genome?

Strictly speaking the question is from the realm of probability theory. If the probability of a nucleotide $X$ in the genome is $p_X$ ($\sum_x p_X=1$), then the probability to encounter a particular sequence, where nucleotide $X$ occurs $n_X$ times, is
$$
\prod_X p_X^{n_X} = p_A^{n_A}p_C^{n_C}p_G^{n_G}p_T^{n_T}.
$$
The formula given in the question is a particular case for $p_C=p_G=g/2$, $p_A=p_T=(1-g)/2$,
$$
\left(\frac{g}{2}\right)^{n_C+n_G}\left(\frac{1-g}{2}\right)^{n_A + n_T}
$$
which is obviously always the case for DNA, where the numbers of Cs and Gs (or As and Ts) are always the same, but the more general formula could serve for RNA or proteins.
What one should keep in mind is that the formula above is for a sequence with a particular order of the nucleotides! If the order is immaterial, and only the content of the sequence matters, the formula has to be multiplied by a binomial factor, which is effectively summing over all the possible arrangements of the nucleotides. Thus, for the case of DNA we obtain
$$
{n_A + n_C + n_G + n_T \choose n_C + n_G}
\left(\frac{g}{2}\right)^{n_C+n_G}\left(\frac{1-g}{2}\right)^{n_A + n_T}
$$
You can check yourself that the difference due to the binomial factor is striking!


Why do bacteria use formylated methionine in the initiator tRNA, while eukaryotes do not?

Summary

The question is posed in terms which imply that the formylation of the initiating methionine (Met) of protein synthesis was a relatively late evolutionary development for some purpose not concerned with translation, such as allowing regulation of protein degradation.

My answer takes the quite different view that this was an early evolutionary development in translation, arising as part of the system for initiating at a specific codon — AUG. With time, the initiating system became more sophisticated, with the specificity of initiation residing more in a separate initiating Met-accepting tRNAf which evolved to have a distinct structure from the elongating Met-accepting tRNAm allowing it to be distinguished from the latter and other elongator tRNAs.

I postulate that in the line leading to eukaryotes and archaea a mutation subsequently occurred that resulted in a loss of ability to formylate Met-tRNAf, but that individuals with this mutation were viable as initiation had ceased to be dependent on the formylation for discrimination.

Contemporary systems for initiating translation

I constructed the diagram below for an answer to another question, but it shows some of the common features of initiation of protein synthesis in contemporary eukaryotes and prokaryotes. (The reader who wishes more detail on translation is directed to standard text books, e.g. chapter 29 of Berg et al.)

The common features of initiation in eukaryotes and prokaryotes are:
1. The use of  a Met-accepting tRNA with an anticodon that recognises AUG to insert the first amino acid in a protein.
2. That this tRNA, here designated tRNAf (but also designated tRNAi) is distinct from the Met-accepting tRNA used for inserting Met at other points in the protein during elongation.
3. That the (f)Met- tRNAf is recognized by an initiation tRNA-binding factor (I-TBF in the diagram — a non-standard abbreviation) and not by the elongation tRNA-binding factor (E-TBF in the diagram).
4. That the (f)Met- tRNAf is delivered to the P site of the ribosome (otherwise occupied by the growing peptidyl-tRNA), rather than the A site, to which elongator-tRNAs are bound.
5. In many cases the N-terminal Met is removed by proteolysis after translation.

The features of initiation that differ between eukaryotes and prokaryotes are:
1. That the initiating amino acid in eubacteria and mitochondria and chloroplasts  is N-formyl-methionine (fMet), rather than methionine, as is the case in eukaryotes and archaea.
2. The selection of the particular AUG codon for initiation is different in the two cases. In eubacteria and archaea the 16S rRNA binds to a polypurine, Shine and Dalgarno, sequence; in eukaryotes the normal method involves a complex of the small ribosomal subunit and the Met-tRNA ‘scanning’ from the 5ʹ-end of the mRNA (the secondary structure of which is first unwound.

Argument

As the question about the function of the formylation is posed in evolutionary terms, and fMet-tRNA is involved in the initiation of protein synthesis, it seems valid to consider the evolution of this latter process, however speculative that may inevitably be. The following formulation is my own, although I suspect it represents the orthodoxy in this field. This, of course, does not mean that it is necessarily correct.

It seems likely that initially translation lacked signals for start and stop (and probably also protein initiation factors). I will not deal with termination, other than to say it is probably an easier problem, but initiation would seem to require the introduction of two new features:
(i) The designation of a specific initiation codon AUG†, which, for whatever reason, corresponded to one of the existing amino acids, methionine.
(ii) Given that this initiation codon also served for incorporation of Met internally, the development of a system to select one particular AUG for initiation.
It seems simpler to suppose that (i) preceded (ii), and that is what I shall assume, although the following arguments do not depend on this.
In contemporary initiation the problem of competition between initiation and elongation factors for (f)Met-tRNAs has been solved by the evolution of two separate tRNAs that accept Met — one for initiation and one for elongation. These share certain structural features — their amino acylation is catalysed by the same amino acyl-tRNA synthetase — but they differ in recognition by the protein synthesis factors that bring them to the appropriate P or A site on the ribosome. Despite the lack of formylation of Met-tRNAf in eukaryotes, the structural similarity to the eubacterial tRNAf is such that it can be methylated by E. coli transformylase (reviewed here). However, I imagine that this structurally different tRNA took time to evolve, as, indeed did the protein synthesis factors.

My argument is that the formylation of Met-tRNA was the basis of the initial discrimination between initiator and elongator Met-tRNAs, and this could have arisen even before the gene duplication that led to separate tRNAs.

Why would fMet-tRNA bind preferentially to the P site and Met-tRNA to the A site? The amino acyl tRNAs that bind at the A site all have a free α-amino group that attacks the linkage of the peptide to the tRNA in the peptidyl-tRNA in the A-site at the peptidyl transferase centre. The peptidyl-tRNA, of course, lacks such a free α-amino group. In this respect fMet-tRNA, with a blocked α-amino group, resembles peptidyl-tRNA (see diagram below). One can imagine how structural differences in the P site could hinder binding of Met-tRNA, but allow binding of fMet-tRNA. And, of course, fMet-tRNA would not be able to react if bound at the A site, which might also have structural features to hinder its binding.

If subsequently the discrimination became magnified by gene duplication and evolution of a structurally distinct tRNAf, the need for formylation would be diminished, and one could envisage that its subsequent loss in eukaryotes may have had no adverse effects, or was easy to accommodate.
The question does arise why formylation has persisted in eubacteria, even though it is possible to obtain viable mutants lacking the transformylase. This is what the work alluded to in the question addresses. There is a 2015 paper by Piatkov et al. which suggests that N-terminal fMet can be a degradation system in E. coli. Presumably this arose after the divergence of the line leading to eukaryotes and archaea, which developed other systems for regulating protein turnover.
However I would suggest that, assuming that the foregoing is correct, it is an explanation for the persistence of the formylation of Met in eubacteria, and not the reason for its initial appearance.

Concluding Remarks

At present I know of no convincing evidence that proves my arguments and rejects the proteolysis idea regarding the reason for the evolution of fMet-tRNA. This is the problem with ideas about early evolution. It is compounded here by the facts that suggestion of a role in proteolysis only came up recently and that this is no longer the ‘hot’ field it was 50 years ago.

Nevertheless, I find it hard to imagine a system as crucial and sensitive as protein synthesis incorporating a feature like formylation of methionine if it only had a secondary function. In this case it would seem easier to formylate the N-termini of proteins post-translationally, as for other N-terminal modifications like acetylation. Furthermore, the finding that the eubacteria-derived mitochondria still use fMet can hardly be to control protein turnover, as the vast majority of mitochondrial proteins are translated in the cytoplasm and lack fMet.

†Footnote

I am aware that, in bacteria especially, there are other possible initiation codons, notably GUG, although the initiating amino acid is always (f)Met. (This attests to the importance of the tRNA structure in contemporary initiation.) It is simpler just to refer to AUG in the argument.


To repeat @Malijam comment, by thermal motion it is referring to the "random giggling" of molecules.

Temperature difference is nothing but a change in the frequency at which molecule move randomly. This movement has no specific direction. If a given molecule is prevented from moving into one specific direction (due to attachement to another molecule) but allowed to move into another, then the molecule will move directionally pushed by nothing but its thermal kinetic. This create a so called thermal ratchet.


In the absence of someone more qualified, I believe this to be a prairie crayfish, based on where it was found (land) and what it is (decapod of some kind that looks like a crayfish). Therefore it is unfortunately not an insect.

It's in the right range and roughly the right terrain.

I welcome entomologically qualified advice!


Is this a simple Hardy-Weinberg problem?

Too long for comment :

If you want to solve the question by Hardy Weinberg, there are certain assumptions involved :-

1.There is random mating. People do not choose their mates on the basis of whether they carry PKU gene or not & mating among close relatives is not there.

2.No new PKU mutations happen.

3.There is no difference in the reproductive capacity of people with or without PKU gene.

If your study population fulfills these criterias, you can apply Hardy-Weinberg to calculate frequencies of various genotypes.

Because this question is  homework ,such assumptions are not mentioned. And, at undergrad level,there is no other concept by which you can solve this question except HWE.

PS: Try to think what would happen if these assumptions are not fulfilled.

PSS: Even if you are solving a small homework question, do think about the logic behind every step. Though it may seem like a lot of work initially , it has enormous potential to teach you subtle concepts !


What species have had their genomes sequenced/are being sequenced?

The GOLD database (Genomes Online DB) contains data on the sequencing status, and also some stats (number of chromosomes, genome size) -- but this extra data is not available for all species.


Are there baby drinks closer to breast-milk?

It is definitely possible to include antibodies and other biological molecules in formula to better approximate that of its mother's milk.  However, there's no way you could mass-produce this. Every mother has unique antibodies, even horomones that are the same in different people are present at different concentrations. The current "formula" on the market is good for as many people as possible.

Isolating antibodies from a mother to include in formula milk would be insanely, insanely expensive. Think thousands of dollars for a carton of formula. Not to mention the hassle of frequent blood and serum draws from the mother for antibody isolation.

So, yes, this is possible (and I wouldn't be surprised if someone with a lot of money to burn has tried it). It's not remotely cost-effective or feasible, though.


(how) is it possible to choose the biological sex of a child?

The main technique is sperm separation or sorting, where you separate sperm by gender, and only use the gender that you want. Existing techniques aren't perfect. AFAIK, they are typically less than 90% accurate, but it is still better than 50%. Typically, this involves some type of cell sorting device often with some kind of fluorescent tag marker.

The second technique is embryo screening. Often, with IVF, you screen and can confirm the gender that you want before implanting the embryo in the mother.

Links:

http://www.fertility-docs.com/programs-and-services/gender-selection/select-the-gender-of-your-baby-using-pgd.php

http://www.in-gender.com/Gender-Selection/MicroSort/


How are blood vessels affected by UV and infrared radiation?

UV rays cause damage is by thinning the walls of surface blood vessels, leading to bruising, bleeding, and the appearance of blood vessels through the skin. Longwave UV radiation (UV-A) accounts for up to 95% of the UV radiation that reaches the earth's surface. Although UV-A is less intense than UV-B, it is more prevalent and can penetrate deeper into the skin layers, affecting the connective tissue and blood vessels, which results in premature aging - Photoaging.

At the same time UVB – is even more dangerous than UVA radiation and also causes skin damage and skin cancer. It affects the surface skin layer. The skin responds by releasing chemicals that dilate blood vessels. This causes fluid leakage and inflammation – better known as sunburn.

I am not quite sure what chemicals are generated by UV radiations beneath the skin surface. I guess it might be excess free radicals which accumulate and cause damage to blood vessels since they are delicate. There is one theory which talks about damages to skin due to Free Radical theory of aging

To give an example how UV radiation affects the blood vessel here:

  In actinic purpura, UV radiation damages the structural collagen that supports the walls of the skin's tiny blood vessels. Particularly in older people, this collagen damage makes blood vessels more fragile and more likely to rupture following a slight impact.

Below figure to give an idea of how strong UV radiations are:


Does using CRISPR/Cas9 knockout need to add donor DNA in the process?

Short Answer:
No, for CRISPR/Cas9 knockout you do not need to add donor DNA.

Little bit more detail:
CRISPR/Cas9 allows you to cut at a given position (defined by the gRNA). This will lead to double strand breaks (DSB).

If you do not add any donor DNA, the cell's DNA repair mechanism will try to repair the double strand break. Since these CRISPR/Cas9 mediated DSBs usually have a blunt end it is really hard for the DNA repair mechanism to fix it. If will often introduce an error, for example by missing a basepair (or several) leading to a frame-shift mutation. This DNA repair mechanism is called Non-Homologous End Joining.

If you do add donor DNA it will contain both the sequence you want to introduce (say a green fluorescent protein) and homologous sequences on both sides of the double strand break. In other words, if you want to cut the sequence ATCGCA exactly in the middle you will get ATC and GCA. You want to insert AGA. The donor plasmid must then have the sequence ATCAGAGCA. As the cell now has the homologous regions, it will use the Homology Directed Repair mechanism.

The sequences are for illustration only - in reality a gene is usually several hundred basepairs and it is recommended to have much longer homology regions.


Small yellow bug with black head and three black stripes on the back

It looks like a yellower version of the brown marmorated stink bug (halyomorpha halys) nymphs I've seen in my garden in the US (west Texas). They change colors throughout their life span, so perhaps that's an older nymph that's begun changing to brown. They're prevalent in the US, Europe, and Asia, according to Wikipedia (https://en.wikipedia.org/wiki/Brown_marmorated_stink_bug).

I have no research experience with this insect, and I am not a biologist -- all of my research on this consisted of google searches for variations on "small red beetle three black lines."


Can radiation equilevant to 1 CT (computer tomography) scan causes significant changes in human body?

As a rule of thumb you can say that a CT scan exposes you to a higher dose than a X-ray examination, due to the different techniques. For CT scans a number of single 2D photos is processed into the 3D models. However, the dose strongly depends on what body part is examined. See a part of this table (the complete table can be found in reference 1):

The additional risk for most examinations is low to very low and since we don't get x-rayed on a daily basis this is a risk which can be tolerated. Besides this, there are other exposures to radiation which also accumulate, but where people usually don't hink about it. These are for example smoking and flights at great altitudes. See the answer from @anongoodnurse for some more details on this.

References:

Radiation Dose in X-Ray and CT Exams
The 2007 Recommendations of the International Commission on
Radiological Protection
AAPM Response in Regards to CT Radiation Dose and its Effects


To understand this, you have to understand how an agarose gel is able to effect the size separation of DNA in the first place.

The reason agarose gels work to separate DNA based on size is that the electric field produced the electrophoresis machine is imparting force on the DNA. The effect of the increasing charge for DNAs with increasing nucleotides is offset almost exactly by the increasing mass of those larger DNA molecules, so in the absence of the gel, all pieces of DNA would migrate through the apparatus at a constant rate.

The reason they don't is because of the mesh framework of the agarose gel. This is like a dense forest or a large number of sieves with random sized holes. There's small holes (of varying size) and in order for large pieces of DNA to work its way through the gel, it has to weave through these pores. Smaller pieces of DNA can go through smaller pores, so they take a faster, more direct path, while larger pieces have to make detours, and even when they find a pore they can fit through have to spend a bunch of time flopping around before they can stuff themselves through the pore.

So it's not strictly speaking "size" or "compactness" by which the agarose gel separates molecules, but instead it's the ability of the piece of DNA to navigate through the mesh of agarose molecules. While relaxed, circular  DNA is more compact than an extended linear DNA of the same nt length, that doesn't necessarily mean that it has an easier time working its way through the network.

For example, a linear piece of DNA can orient itself so it goes "end first" through the mesh. Once the end of the DNA goes through a pore, it's relatively easy to pull the rest of the DNA through the same pore, as compared to trying to bend the DNA and pull it "sideways" through the pore - you have to remember that on molecular scales, DNA is rather stiff. You can't kink it in half very easily, so a piece of DNA that has to go through the pore "sideways" takes much more space than one that can be threaded through end-wise, even if you bend it as much as possible into a hairpin.

With a relaxed circular DNA, there's no end, so it's always going through the pores "sideways" so it's limited to the larger pores. In addition, it's large and floppy, so it takes quite a while to go through the pores that are there. When you add supercoiling, you don't fix the issue with going through pores "sideways", but you do limit the flopping around it does, so once you do find a pore of a usable size, you get the rest of the DNA through faster. In fact, with enough supercoiling you reduce the time spent flopping around enough that it goes through the agarose even faster than the linear DNA, which - while it can fit through smaller pores - still spends a bunch of time flopping around instead of going through the pore that it's started to go through.


What do the acronyms in C.elegans neuron names stand for?

Unfortunately, there doesn't seem to be a very systematic nomenclature. Some of the letters are acronyms while others describe a class of neurons. The letters V and D usually describe the ventral and dorsal counterpart of a pair of neurons. The same goes for L and R (left and right).

This list names all C. elegans neurons and you can see that, for example, the "ring interneurons" or motor neurons start with RI or RM. "Inner labial" or "outer labial" neurons are called IL or OL. Ventral cord motor neurons start with a V and their different classes are called VA, VB, VC and VD plus a number. The classes are determined by lineage origins or historic reasons ;) And there are a lot of exceptions to these rules, because their historic names remain even though we continue to learn more about these neurons.

The example AVAL/AVAR are ventral cord interneurons. They form a class with other ventral cord interneurons called AVB, AVD, AVE. So, the first two letters refer to the main class, the third letter to the subclass. The L or R refer to the left and right member of this pair.

Some friends working in C. elegans neuroscience told me that you basically just treat these letter codes as individual names for the neurons without bothering about their origin and meaning too much.


I don't think it's reasonable to try to do this in a species-independent manner, since there is so much difference between ant species. Leaf-cutter ants, for example, will build trails to forage 200 meters away from their nests. Likewise, ants vary by two orders of magnitude in size and the walking speed of an ant will vary accordingly as well. Finally, I don't think that you can reasonably exclude trail-building from your simulation, since that's a key ingredient of foraging.
Bottom line: pick a species to model and look for information on its specific mobility.


The pollex was a Roman length measurement, approximately equivalent to an inch. The pollex was also known as the uncia, which is where the word "inch" comes from. There were 12 unciae in a pes ...


Can a ladybug be determined just by its number of points?

No, most of the time you cannot. There is a large diversity of ladybug species (several dozens in western Europe), and many of them can have the same number of spots. Moreover, as you already said, there can be intraspecific variation in the number of spots.

I do not know if there are specific numbers of spots that would be associated to only one species - but for most numbers, it is not the case. For instance, yes, it is common for Harmonia axyrids to have 18 spots (source). But Anisosticta novemdecimpunctata, which  can also be found in Europe - and can also be orange ! - may have between 15 and 21 spots (though it more frequently has 19) (source).

Thus, the number of spots cannot be used as a sole criterion to identify ladybugs: other criteria usually include the general shape, colour of elytra, size, shape and colour of the pronotum, length of antennae, etc. (See here for examples.)

First picture: Harmonia axyridis

Second picture: Anisosticta novemdecimpunctata


How can rhodobacter possess a single 'PS-II' like reaction center?

McEwan (1994) Photosynthetic electron transport and anaerobic metabolism in purple non-sulfur phototrophic bacteria. Antonie Van Leeuwenhoek 66: 151-164.

...The purple non-sulfur bacteria ... carry out photosynthesis under anaerobic conditions. Photosynthetic electron transport is cyclic and involves ... two redox complexes ... reaction center (RC) [and] cytochrome bc1 complex. ... This cycle of electron transfer, [mediated
by ubiquinone (UQ) and a c-type cytochrome,] is linked to the generation of a proton motive force ... across the cytoplasmic membrane.


Hill's function for translational regulation

Your logic looks correct to me. Essentially, what you are doing is uniformly distributing the regulator among the available mRNA.

Note that even when using Hill functions to model transcription, the ratio of transcription factor (TF) concentration to the number of TF binding sites must be large - otherwise, you would have to consider binding ratios even at the level of transcription modeling, similarly to what you are doing now for translation. E.g. consider a hypothetical situation where 10 TF molecules compete for binding to 5 different promoter regions, each having 10 binding site repeats - you need to somehow distribute the available 10 TF molecules among 50 target binding sites. Clearly, this is not something accounted for by the standard Hill equation, which would in such case wrongly assume that 10 TF molecules are regulating each construct.

In your specific case, introducing the ratio of the regulator per each mRNA may not even be necessary if this ratio is large, effectively leading to saturation levels anyway. Note that the maximum level that $m_x$ can reach (at steady state) is equal to $max(m_X) = \frac{\alpha_{mX}}{\beta_{mX}}$. If you can ensure this value to be much smaller than your translational regulator concentration, you will get similar results even if you use simply $\frac{K}{K+R}$ for translation modeling.

If the latter assumption cannot be made, then your reasoning makes sense. For each individual mRNA molecule:

$$\text{X produced per mRNA} = \alpha_{X} .\frac{K}{K+R'}$$

where $R'$ is the amount of mRNA bound to this molecule. If $R$ is the total available regulator concentration and uniform binding affinity is assumed, this means that $R'=\frac{R}{m_X}$. Summing this over a total of $m_X$ mRNA and considering degradation yields exactly your final ODE.

Note that another important assumption you are making is that regulator binding/unbinding to/from mRNA is fast compared to translation, and that no cooperative interactions are present.

If you want to verify things further, you could establish system reactions, then derive Hill-free ODE equations according to the law of mass action. You could then compare this model to the one you propose. Furthermore, performing a stochastic simulation might make sense. If you want to go down this road but don't want to implement Gillespie's algorithm on your own, you can use e.g. SGNSim or COPASI.


Determine the chances of blood group type and rhesus factor of unknown parent?

Simple explanation: that "Blood Type Calculator" is wrong.

Inspecting the source code of that page we can find the if statement that chooses the outcome for the unknown parent, regarding the Rh factor. And it has a problem. Whoever knows JavaScript will understand the problem, but even people that don't know programming languages can follow this, it's not complicated. Let me explain it.

When you choose the Rh blood type in the dropdown menu, the Rh+ blood is associated to the value p and the Rh- blood is associated to the value n. Those are just strings, the programmer can chose any value he/she wants, that's not important, as you know. The variables that hold those values are msel2 for the known parent and csel2 for the child.

Now comes the if statement:

var frh = "+";
if (csel2 == "n" && msel2 == "n") frh = "-";

Let's break it down:

The default value that will be printed on the result, called frh, is + (positive), as you can see before the if statement. As already said, msel holds the value for the known parent, and csel holds the value for the child.

In the if statement, only if the user chooses the known parent type as Rh- (msel2 == "n") AND the child as Rh- as well (csel2 == "n"), the value of frh will be changed to - (the double && represents the AND). Any other combination will not change frh, which will remain +.

And that's plain wrong!

There is no need for me to explain you the genetics of Rh factor, there are plenty of resources online and answers here at Bio SE as well, but here is a summary:

Given that Rh+ individuals can be either DD or Dd, it's easy to show that the unknown parent can be Rh- in case that the known parent and the child have both Rh+:

(Known parent)     (Unknown parent)
    D_ --------------- dd
              |
              |
             Dd
           (Child)

The lesson to learn here: don't trust everything you find online.

Correct algorithm:

For your second task ("Determine the possible blood group and rhesus factor of an "unknown parent" based on the blood group and rhesus factors of the child and the known parent."), let's see all possible cases:

Known parent Rh+ and child Rh+: the unknown parent can be either Rh+ or Rh-.
Known parent Rh+ and child Rh-: the unknown parent can be either Rh+ or Rh-.
Known parent Rh- and child Rh-: the unknown parent can be either Rh+ or Rh-.
Known parent Rh- and child Rh+: the unknown parent has to be Rh+.

As you can see, in only one case the unknown parent cannot be either Rh+ or Rh-. That being said, this is a possible solution:

var frh = "+/-";
if (csel2 == "p" && msel2 == "n") frh = "+";

What this snippet does is this:

It assigns the default value for the unknown parent (frh) as +/- (meaning that he/she can be Rh+ or Rh-, we can't know) for all possible combinations. However, if the mother is Rh- AND the child is Rh+, that value is changed for + (because, in that situation, the unknown parent has to be Rh+).


What is the Baculovirus and how does it help in research?

Baculoviruses are a a class of viruses which infect only a pretty small number of insects (or their cells in cell culture). To do so, the virus has to infect a cell, take it over to produce the own proteins and DNA to make new virus particles and finally set these new particles free. This is basically what is shown in the image below (from here):

From the proteins encoded by the viral genome, not all are essential, meaning removing them will not do harm to the virus or its function. And this is used in Biotech to produce proteins of choice. One of the non-essential proteins is replaced by the sequence of the protein you want to have produced by the cells. You then take insect cells in cell culture (these are single insect cells which live in a big vessel full of culture medium), infect these cells with the modified virus and eventually harvest the protein from the culture.

The advantage of Baculoviruses is that they only infect the insect cells on which they are specialized so they are a safe way of producing proteins for pharmaceutic applications. If you want to read some more about this, start here.


Is half-a-tablet of X a substitute for a tablet of X/2?

This website gives a nice overview, including how the FDA approve drugs that they expect will be spit and some practical issues of pill cutting.

Yes, half a tablet of (2x dose) is, absent other issues, equivalent to one intact tablet of (1x) dose.  The main exceptions are practical ones.. where the tablet cannot easily be cut; where the dosing is so specific that slight variations of dose (uneven tablet fragments) matter, or where the dose is in a capsule or has some special coating so that cleavage might alter the release of drug into the body.

Yes, dosing scales upwards.  Two pills give twice the total dose of one pill.

All prescriptions are dispensed with specific dosing regimens (i.e how much you take and when you take them) to ensure the drug reaches effective concentrations in the body. You should not try to 'boost' the drugs effectiveness by taking more drug, or taking drug more frequently than your physician has prescribed.  As noted in the weblink above, it can be tricky to keep track of dosing and timing if your pill bottle contains whole and cut pills.  (i.e what does "take one pill every NN hours" mean ?).

This is NOT MEDICAL ADVICE . Always consult your physician and/or pharmacist for your particular case.


Could plants do photosynthesis at moonlight?

Of course they can and do, except in total darkness (spectroscopically, only bands in the far red and in the blue spectra matter - blanking these affects 'total darkness').

In photosynhesis a photon is adsorbed by Photosystem II to break down water into oxygen and protons in solution. Another photon must be adsorbed by Photosystem ! to power the enzymatic machinery that make NADPH and ATP that power the Calvin cycle.

We know the sun powers photosynthesis effectively. The sun produces somewhat less than 100,000 lux (lumen per square meter). Moonlight is one one millionth of this or about 0.1 lux. One lux is something like 10^15 photons per second (per square meter); so moonlight provides something approaching 10^14 photons per second to drive photosynthesis. The spectrum of moonlight is not markedly different from that of sunlight. So, moonlight provides an ample number of photons per second per square meter to power photosynthesis.

The trouble, though, is that the rate of photosynthesis is low compared to the rate of metabolism in the rest of the plant. So, in effect, the plant gives off carbon dioxide by night and oxygen by day even though both gasses are being emitted all the time.

EDIT (per request) some references:

Photosynthesis
Respiration
Moonlight lux versus sunlight
Photons per second in a lumen


Why do diseases in the tap water of developing countries affect people from developed countries more?

You can expect that someone drinking from a water source regularly will develop immunity to the pathogens in that water source by being repeatedly exposed to them.
However, it's not quite right to say that poor quality drinking water impacts mostly tourists. It mostly impacts the people drinking that water, especially children. Immunity comes at a cost. Some example statistics from Wikipedia:

Between 2000 and 2003, 769,000 children under five years old in sub-Saharan Africa died each year from diarrheal diseases

In South Asia, 683,000 children under five years old died each year from diarrheal disease from 2000 to 2003. During the same period, in developed countries, 700 children under five years old died from diarrheal disease.

And from WHO:

Contaminated water can transmit diseases such diarrhoea, cholera, dysentery, typhoid, and polio. Contaminated drinking water is estimated to cause 485 000 diarrhoeal deaths each year.

These statistics conflict a bit, and I won't go too far into the reasons why or try to identify the correct number. I think they are all sufficient to identify this as a substantial issue in the world.
Responses to disease vary between individuals for all sorts of reasons, including genetics, overall health at the time (for example, someone well-nourished versus poorly nourished), and access to medical care and treatment. Some you could also add up as effectively "luck". In a hypothetical scenario, you could expose 100 people to a pathogen, and of those 100, many may get very sick and a few die. The survivors may then have future immunity to the same pathogen, but it would be wrong to say the population as a whole is not affected by it, you're just looking at the people who survived.


How does hyperthyroidism cause diarrhea?

Hyperthyroidism leads to an overactive sympathetic nervous system, hypermotility and diarrhea (Culp and Piziak 1986) (Geffner and Hershman 1992) (Maitra 2015, p. 1084) (Thomas et al. 1973). A demonstration of this is the patient presenting with the sole symptom of intractable diarrhea. It was found to be secondary to hyperthyroidism, and was treated successfully with β-adrenergic antagonists (Bricker et al. 2001).

Was the hypermotility due to excessive β-adrenergic activity, or insufficient α-adrenergic activity?

An article by Silva and Bianco (2008) states that β-receptor expression is under transcriptional control of a thyroid response element, and is upregulated by increased thyroid signalling. There is much less research on α-receptor expression, I would be much obliged if anyone can provide me with access to the following article:

  *Bilezikian, JP, Loeb JN 1983, ‘The Influence of Hyperthyroidism and Hypothyroidism on α and β-Adrenergic Receptor Systems and Adrenergic
  Responsiveness’, Endocrine Reviews, vol. 4, no. 4, pp. 378-388.*

In any case, they add that increased β-receptor expression is rather modest, often overestimated, and cannot account for the magnitude of amplification in responsiveness induced by thyroid hormone (TH). Rather, thyroid signalling increases cAMP levels associated with a2 and β-receptors to a degree which could account for these changes.

Increased cAMP occurs during normal β-receptor function but is significantly potentiated by TH in several ways: increases in adenylyl cyclase; downregulation of β-receptor G-protein subunits, Gai and Gβ; and downregulation of phosphodiesterases which degrade cAMP. This explains the effectiveness of the β-adrenergic antagonist noted above, as propranolol decreases cAMP levels, thus decreasing hypersensitivity of the hyperproliferated β-receptor (Silva and Bianco 2008).

  “The reduction of the sympathetic activity associated with
  hyperthyroidism outflow (via increased rate of norepinephrine
  metabolism to decrease overall levels) is likely overridden by the
  increased sensitivity, as suggested by the clinically significant
  amelioration of the adrenergic symptoms with β-adrenergic blockage and
  the consistent observation of elevated cAMP production.” (Silva and
  Bianco 2008).

What are the effects of preferential/hyperactive β-receptor function on gastrointestinal motility?

The β1-receptor increases ghrelin secretion from the stomach – a peptide secreted when the stomach is empty to increase hunger, increase gastric acid secretion and gastrointestinal motility. The β2-receptor promotes smooth muscle relaxation (decreased GI motility) but contracts sphincters of the GI tract (Adrenergic receptor 2018).

β-adrenergics also promote the secretion of gastrin – a peptide which increases antral muscle mobility and promotes stomach contractions; relaxes the pyloric sphincter, which increases the rate of gastric emptying; is involved in the relaxation of the ileocecal valve; induces pancreatic secretions and gallbladder emptying; and contributes to the gastrocolic reflex (Gastrin 2018) (Katzung and Trevor 2014, p. 143-144).

What are the effects of relative α-receptor deficiency on gastrointestinal motility?

Considering that β-receptor antagonism causes a relative increase in α-receptor activation we should consider them also. The α1-receptor causes smooth muscle contraction. The α2-receptor causes negative feedback on norepinephrine signalling, relaxes the gastrointestinal tract when activated on presynaptic terminals and contracts of sphincters in the GI tract. It does this by decreasing levels of cAMP (whereas β-receptors work by increasing cAMP) and so it will become less active as TH becomes more active (Adrenergic receptor 2018) (Silva and Bianco 2008).

  What are the adrenergic-mediated effects of hyperthyroidism on gastrointestinal motility?

  Under prolonged conditions of excess TH overall peristalsis will
  increase due to mixed effects on smooth muscle contraction primarily
  via α1, and β2. There will be an increase in hunger and gastric acid
  secretion via ghrelin (Adrenergic receptor 2018). Hyperphagia and
  hypermotility are both suggested to play a part in steatorrhea
  (Culp and Piziak 1986).

How does hypermotility cause diarrhoea?

In addition to the mechanisms above, it has also been suggested that excessive fatty food intake contributes to the steatorrhea seen in patients with hyperthyroidism. This is compounded by a reduction in time that digestive secretions can act on chyme, ultimately leading to a reduction in absorption and an increased liquidity of the stool (R Santra, Calcutta School of Tropical Medicine, 2016).

Finally, it is important to note at this point that not everyone with hyperthyroidism will show hypermotility - some hyperthyroid patients show delayed gastric emptying (Pustorino et al. 2004). Indeed, constipation has been reported in patients with hyperthyroidism (Culp and Piziak 1986). Nonetheless, it is held that dysthyroidisms alter the motility of the gastrointestinal tract, and we do not completely understand how this works (Pustorino et al. 2004).


Do you have nerves on the outside of your skull?

There are a number of nuerovascular bundles that exit the skull. Three involve branches of the trigeminal nerve.


I've found that extracted RNA using commercial kits has stayed stable for many years at -80 C. I would certainly aliquot it before freezing however as RNA is particularly sensitive to freeze-thaw cleavage.


Why are genes expressed as proteins rather than other types of bio-molecules?

Wow - like a biophilosophy question! The answer lies in in building up the largest diversity with minimal subunits. Efficiency if you will. In this case we are talking about amino acids. We only need 20 of them to approach unfathomable numbers of diversity.

The protein acts as a messenger, as an enzyme, a carrier and it's own building unit. If you wanted that kind of range out of any other known biomolecules, you would need a gene to code for each of them, and itself a gene a completely different mechanism to build them which we can't even fathom. It's so much simpler to code for a protein, which can act as an enzyme to build other biomolecules.

However if you are asking about why the central dogma of molecular biology is the way it is...well then, it seems to be axiomatic.


This due to a phenomenon called "cold shock". This induces a number of physiological changes in the fishs metabolism and also in its behaviour and can lead to death.

The first paper cites some reasons in table 1:

Brain and central nervous system response: Changes in neuronal
activity
Catecholamine and corticosteroid response: Release of hormones due to
the shock.
Haematological and metabolic response: Changes in red and white blood
cells and lactic acid in the blood.
Protein expression and molecular responses: Changes in the expression
of proteins of the heat shock family.

These changes (especially in brain and nerves and also the hormones) can deregulate the fishs metabolism so much that it can die. Shock is a critical condition. Obviously a lot of research is going on, but I recommend reading the two articles below for further details.

References:

Cold shock and fish
Physiological stress responses in the warm-water fish matrinxã
(Brycon amazonicus ) subjected to a sudden cold shock


No, I don't think it looks like Heracleum sosnowskyi. To me, the plant in your pictures looks similar to Angelica sylvestris (which is common in Poland), but there are many species in Apiaceae with a similar apparence and I'm not familiar with all of them. Species in the Heracleum genus has leafs that are "lobed" and all leaflets aren't as separated as the plant in your pictures (overall, the leafs of Heracleum look quite different), and the stem is usually "swollen" where the leafs connect (i.e. the leaf stems are swollen). Plants of Heracleum sosnowskyi (and H. mantegazzianum) are also much larger, and they often have a fussy apparence because of the bristle hairs.

Angelica sylvestris (picture from from wikimedia commons)


In men, gout is associated with a higher risk of death from all causes. This would imply that their life expectancy is shorter. From a review by Kim et al. (1):

  Among men who did not have pre-existing coronary heart disease, the
  increased mortality risk is due primarily to an elevated risk of
  cardiovascular death, particularly from coronary heart disease. Also,
  an extension study of a large clinical trial among men with
  above-average risk for coronary heart disease found that a diagnosis
  of gout accompanied by an elevated uric acid level is associated with
  increased long-term (approximately 17 years) risk of all-cause
  mortality that arises largely from an increased risk of cardiovascular
  disease (CVD) mortality.

This does not rule out that some subset of people with gout (patients with less severe symptoms?) might benefit in some way from an antioxidant effect, but in general gout is obviously not good for life expectancy.


It looks like certain species of earthworms need to surface because they have higher oxygen consumption, while other species do not need to surface because of their lower consumption.

In particular, specimens of Amynthas gracilis were found to have low water tolerance, hence they

  crawl out of the waterlogged soil, especially at night when their oxygen consumption increases

while Pontoscolex corethrurus

  survived longer under water and was never observed to crawl out of the
  soil after heavy rain

Role of diurnal rhythm of oxygen consumption in emergence from soil at night after heavy rain by earthworms


Is chloroplasts per cell indicative of the photosynthetic power of a plant?

You can say that but chloroplasts do not have uniform morphology across different species. Moreover some organisms such as red algae have chloroplasts of different origin.

Real indicator would be lets say number of chloroplast ATP synthases and/or light harvesting photosystems; these can be perhaps indirectly approximated by the total surface area of the thylakoids.

But for all practical purposes and comparison between not so distant species density of chloroplasts is a decent enough approximate for photosynthetic power.

About turning carbon dioxide to oxygen:

You should note that the source of oxygen is not CO2. Oxygen comes from photolysis of water; this is the initial step in the electron transport chain (ETC) in chloroplasts. The ETC results in production of ATP and NADPH; the latter is used in the Calvin-Benson cycle to fix CO2 to sugar.

So photosynthesis and carbon fixation, though associated in chloroplasts are basically different biochemical processes. Theoretically as long as NADPH is provided a plant should be able to fix CO2 [this is just my guess].

It is difficult to observe internal structure of chloroplasts with normal bright-field microscope but you can count them easily at 40x magnification. See here.


How would one go about isolating a specific mRNA out of a pool of others?

First of all you need to figure out what your goal is: an expression vector (i.e. plasmid) which will drive expression of a gene in bacteria (let's say E.coli), and your gene's coding sequence needs to be downstream of the promoter in that construct.

Your task is asking for a pretty routine lab task known as cloning (https://en.wikipedia.org/wiki/Molecular_cloning). Your mRNA pool might be the result of an RNA extraction from a tissue sample and your goal is to get bacteria that produce lots of the desired protein.

As this is homework, I will only give a broad overview and leave you to figure out the details.

Indeed one way you could approach this is actually some method to specifically extract the desired mRNA, then generating cDNA from it and pasting that into the vector. This is unlikely to give you good success rates, and in practice nobody purifies a specific mRNA unless they are interested in other molecules naturally bound to that mRNA.

Since this is a procedure with a very practical purpose without a scientific question behind it, there's a much more efficient way to achieve it.

Generate cDNA for all mRNAs in the mix ("first-strand cDNA synthesis").
Instead of removing unwanted RNAs, simply amplify the desired one using specific primers you designed and PCR. This gives you what is called the "insert".
Most likely if you ordered the expression vector online, you actually receive bacteria containing it, so you first have to culture those and extract DNA from the culture ("miniprep").
Next step is to paste the DNA into the expression vector. For this you need to digest both the vector and your insert using the same restriction enzymes ("restriction digest"). (Your insert will not have restriction sites on it, so you will need to introduce them by designing the primers for the PCR in step 2 appropriately.)
Run the restriction digest mixes on a gel (electrophoresis), identify the bands corresponding to the insert in one lane and the vector in the other, and extract them (gel extraction).
Ligate the insert and vector
Transform the finished product into competent E.coli


Can viruses protect themselves against restriction enzymes?

I guess you are talking about the restriction modification system, not restriction enzymes in general (which are used a lot in the lab, for example).
If so, this paper might help answer your question.

I have not read through it totally, but at least some viruses trigger the downregulation of restriction enzymes to make them cleaving their DNA less likely. Restriction enyzmes are also quite selective for the site they cut at, so single nucleotide changes at this site, might, if the phage is lucky, not change the function of the encoded gene, but keep the restriction enzyme from binding (and thus cleaving) at that site. Mutating helps a lot. And it is fast in those little critters.


In microarray normalization, why is the normalization factor this?

$G_k^{'}$ and $R_k^{'}$ are normalized values of $G_k$ and $R_k$.

Take say G as $[1,2,3,4]$ and R as $[100,150,200,400]$ as your values and you want to normalize them. This is scaling one of them onto the other and bringing them on an equal level to compare. So in your case the factor is $85$ units. So a unit of R amounts to $85$ units in G.

So to scale G to R, multiply G by $85$ or you can scale R to the level of G by dividing R by $85$. So the values are either $[85,170,..]$ and $[100,150,200,400]$ or $[1,2,3,4]$ and $[1.17,1.76,..]$ from our example.

I think it should be mentioned as $T^{'} = R_i/(G_i*N_{total})$ as in log ratio from the statement in your question.


What is the advantage to neurons being asymmetrical?

Asymmetry to essential to neuronal function, particularly in the peripheral nervous system. This is because the dendrites of a neuron receive synaptic potentials, which are graded and decay with distance. Dendrites are typically short such that a synaptic potential is still above threshold by the time it reaches the axon hillock (sometimes called trigger zone), so that a subsequent action potential can fire. Axons need to be long (some are up to a meter long, in the leg) to transmit signals long distances in the body. To make a neuron symmetrical, one of these qualities would have to be compromised and neuronal function would suffer as a result.

Source: Principles of Neural Science, 5th ed (2012), Kandel et al. Chapters 1-6.


Do humans perceive temperature or heat-flux? (or both?)

This is the modified answer in response to the discussion:

Facts:

There are warmth and cold receptors in the body at two places: The Peripheral receptors and the Central Receptors
The peripheral receptors are present in skin and the central receptors in the body core at multiple sites the notable site being the hypothalamus
The Temperature receptors have two patterns of firing - The tonic response (sustained firing rate due to steady rise and maintainance in temperature over long time) and the phasic response (also called dynamic response - response to short changes in the temperature)
The Warmth receptors fire maximally at 44 - 46 degrees celcius. The firing stops almost completely at about 48-49 degrees celcius beyond which there is no effective thermoregulation
The Cold receptors fire maximally ar 20 - 35 degrees celcius.

What is measured?

The peripheral receptors sense the temperature of skin.
The central receptors measure the core body temperature.

Only the temperature is measured as WYSIWYG mentions.

However the frequency of stimulus (How frequently the nerves fire - this depends on the temperature range in which the respective receptors fire) and the two types of receptors present on the skin allows the brain to interpret the flux. So Chris Stronks is correct in telling the heat flux is perceived.

Mechanism of feeling cold:

When you touch a cold object the heat is lost through conduction and radiation through the portion of skin that is in contact with the object. The cold will cause vasoconstriction of that part of skin - This means that the temperature of that part of skin rapidly equilibriates with that of the object. As there is rapid cooling (assuming that the object is cold) the rate of firing of the warmth receptors decrease and the rate of firing of the cold receptors increase. This allows the brain to interpret that the skin is losing heat (the flux).

The radiation part of heat loss is more appreciated when you pass a uncurtained window during the winter - the temperature difference is so great that very high loss of heat occurs through radiation that you shiver. You have to note that during these instances there is no decrease in the core body temperature. The actual decrease in core body heat needs a long exposure to such conditions. In the scenarios detailed above the heat loss is primarily from skin which reflects the ambient conditions (very quickly). Thus the peripheral receptors effect Anticipatory Control which is effectively measuring the flux of heat through skin.

The opposite occurs when you suddenly stand beneath the sun coming from a cold room - the radiant temperature of the sun being so great will cause the skin temperature to rapidly rise causing the body to feel very hot (though the core body temperature has not changed much).

Interesting details that may increase your understanding (though not directly related to the question):

The heat control effected through sensing the change in the skin temperature is called Anticipatory Control. In this the brain "anticipates" what is going to happen - for example a sudden cold draught - and will respond to it - reponds by increasing the heat production (shivering).

The heat control effected through sensing the core body temperature is Negative Feedback Control. In this the brain responds to increase in the core body temperature and takes steps to remedy this (the response will bring the temperature back to the set point).

For more see:Boron & Boulpaep Textbook of Medical Physiology, 2nd Edition, Chapter 59.


What is the title of Darwin's paper on cellular structure?

Darwin had written lots of papers which dealt with fertilization (which involves single cells), as he primarily dealt with the reproduction, continuation and thus evolution of a species, but these did not deal with cell biology specifically.

After a closer look at the complete list of all his publications, I found that he did have two cell-related publishings, specifically about the size of cells, one entitled Bees in Jamaica increase the size and substance of their cells, and the other, Bee-cells in Jamaica not larger than in England, in which he retracts the statement he had made in the previous article.

Ultimately, to answer your question, it seems as though none of his works dealt with cellular structures.

References

Charles Darwin. "Bees in Jamaica increase the size and substance of their cells". Journal of Horticulture (15 July): 305.
Charles Darwin. "Bee-cells in Jamaica not larger than in England." Journal of Horticulture (22 July): 323.


Method of figuring out how much of a certain mRNA is present in a bacterium

You should be running qPCR to quantitate how much of a certain mRNA sequence is present in a sample. Remember that there are thousands to tens of thousands of different mRNA sequences present in an RNA prep, not to mention ribosomal RNAs, transfer RNAs, etc. If you then run your RNA prep out on a gel, you're just going to get a smear. Even if you cut out a portion of the gel that corresponds to the expected size of your mRNA of interest, there are going to be lots of other sequences present in that portion as well, so doing a gel purification step then measuring concentration with Nanodrop (which is quite accurate within its working range) won't tell you anything.

Quantitative real-time PCR (qPCR), on the other hand, when run with appropriate controls and standards, can give you a very good idea of approximately how many copies of your sequence of interest there are per nanogram of total RNA.


PCR efficiency or DNA yield with single primer

Technically, if you use only 1 primer, it will not be PCR anymore.

But that's what gonna happen if you have 100 template DNA molecules (made of strand A and complimentary strand B) and abundance of primer (say, 10x, or 1000 molecules):

At first cycle primers will bind to template DNA, but only to one strand (for example, A)
DNA polymerase will extend those primers, creating complimentary strands B'. Some strands B' will be shorter than template because DNA pol is not perfect
DNA-primer complex will melt at the beginning of cycle 2
At the beginning of cycle 2 primer molecules will again bind to template, again to strands A, and process continues

Since there is no step where strand A is being copied, this process will be linear, not a "chain reaction". At each cycle you will get the same amount of strands B'.

It is hard to say how much B' molecules you will get at each cycle, but at the end of 30 cycles, it will be at best 30x of the first cycle. Remember, though, that DNA polymerase degrades, and cycle 30 will be less efficient than cycle 1.

Assuming perfect polymerase, and that extension time is long enough, you will get 100 strands B' on cycle 1, 100 strands B' on cycle 2 and so on. At the end of 30 cycles you will have 30x100 strands B', and 100 strands A and 100 strands B.


Leaves have no need to 'exhale' carbon dioxide - it gets consumed by photosynthesis.

Elsewhere in the plant, metabolic processes produce carbon dioxide. This escapes via inter-cellular gas channels that, in woody plants, terminate as lenticels (little pores) on the stem surface. Oxygen for metabolism diffuses into the plant in the same inter-cellular spaces as carbon dioxide diffuses out.

With roots, the oxygen necessary for metabolic processes is dissolved in water.

If you've meant to ask about leaf adsorption of carbon dioxide and when stomata close, it depends on what carbon fixation scheme the plant exhibits, CAM, C4, or C3.

You might want to also consult an  answer I gave to a similar question previously posed here.


How small does a nanobot have to be to "swim through the brain" and access any neuron it wants to?

Is there a visual demonstration I can see of approximately
how "packed full of cells" the brain actually is?

Yes. You can inject tracers in the CSF (Cerebrospinal fluid) or in the ventricles and monitor them. In this study, the authors have injected a fluorescent tracer in the CSF and used a direct imaging.

  We used in vivo two-photon imaging to compare CSF influx into the
  cortex of awake, anesthetized, and sleeping mice. The fluorescent
  tracers were infused into the subarachnoid CSF via a cannula implanted
  in the cisterna magna for real-time assessment of CSF tracer movement.

You can also inject a radiolabeled tracer. The tracer should be a molecule that is not taken up by the cells. You can then visualize the space using Positron Emission Tomography (PET).

 But what is a good ballpark size to be to swim through with relative ease?

You may find this interesting: the abovementioned study shows that the "space" expands during sleep and CSF percolates deeper into the brain. This allows clearing of toxic metabolites. Having said that, some interior regions would be difficult to reach. Apart from neurons, there are several glia which occupy the space. I am not entirely sure about this but in interior regions there would be very less of extracellular space (I am just guessing it would be less than ~1µm. See the black regions in the figure below). Extracellular space is 20% of total brain tissue but it is not constant throughout. For more details refer to this book.


Specific numbers of nucleotides in Okazaki fragments

The length of Okazaki fragments is not necessarily a tight distribution. The lengths are determined by the spacing between adjacent sites where DNA primase has synthesized a short RNA primer on the lagging strand at an active DNA replication fork. In E. coli, as I recall, this occurs on an average of once per 1000 nt.

DNA polymerase holoenzyme then uses that 3'-OH on the RNA primer as a substrate and synthesizes the DNA strand complementary to the Template strand UNTIL it bumps into the next RNA primer.

It is those nicks in the newly synthesized lagging strand, that have a deoxy-3'-OH, and a ribo-5'-phosphate, (that need to be removed and ligated) that mark the boundaries of individual Okazaki fragments.


What is inside IDL (intermediate density lipoproteïn)?

Taken from Fundamentals of Biochemistry (Voet et al. 2013). Note the differences in apolipoproteins between LDL and IDL, as well as the ratio triacylglycerol : cholesteryl esters between VLDL and IDL.


Cell division happens by division at the middle of the rod, so the result is two daughter cells that are at nearly the same angle. Over time in the absence of agitation moving cells around, this will lead to a groups of cells that have non-independent cell orientations.

For more information, including how people have induced it to do otherwise, see this paper.


ATCC- you can get most bacterial strains and cell lines there.

Find P. denitrificans here:

www.atcc.org/products/all/13543.aspx


How do bull semen companies get the semen out of the bulls?

Colorado State University has two helpful pages that explain the main procedures (h/t Tom). Semen Collection goes through the 3 ways that are used to collect semen from animals.

Semen Collection from Bulls helpfully expands on bulls specifically:

  Semen is most commonly collected from bulls in bull studs using an artificial vagina, as described below. Electroejaculation is an alternative method used with bulls that cannot mount or are too fractious for easy handling (e.g. range bulls). Finally, semen can be collected by message of the seminal vesicles and ampullae per rectum.

In a nutshell an Artificial Vagina (AV) uses thermal and mechanical stimulation to stimulate ejaculation. In the image below, the top item is an AV for use with rabbits and the bottom one is an AV for use with bulls. (The middle item is a director cone, which is used to collect semen from dogs.)

The procedure revolves around using a steer as a teaser to arouse and mount the bull, and then having an operator divert the penis the AV:

As one of the pages helpfully explains:

  Prerequisites to use of an AV are that the male be conscious, not significantly frightened of people, and more interested in ejaculating than in killing humans.

The main alternative for less docile bulls appears to be electroejaculation, which involves applying a series of short, low-voltage pulses of current to the pelvic nerves which are involved in the ejaculatory response. You then collect semen using a director cone much like the one in the image further up.


Niacin (better known as vitamin B3) and nicotinamide are both precursors of the coenzymes NAD and NADP in vivo. As you can see below, the biosynthesis of NADP+ proceeds as follows, when nicotinamide combines with a basic ingredient, a sugar phosphate:

The sugar phosphate precursor can also form AMP and GMP, also important molecules in the cell, when it reacts with the nucleobases adenine and guanine (A and G from the 'ladder' that bridges the DNA helix).

Just as an additional funfact, some bacteria such as Plasmodium falciparum and Toxoplasma gondii cannot synthesize nicotinamide themselves, because they lack the genes for it. Instead, they salvage the enzymes from eukaryotic hosts. This is an example of an incomplete biochemical toolkit giving rise to parasitism.


Immunopanning in my experience takes an awful lot of optimization, but is worth it when it works.  I would plan, if you have the resources, to sac rats just for assay optimization before you even begin to try to get real data out (assuming your planning for a steady stream of primary cells).

That said, there are some real easy checks you can try do first.  Try to get a rough cell count on what you put on your negative plate (depending on your prep steps, that can actually be a bit tricky), and then count the cells again as you transfer the cell suspension to the positive plates.  If you don’t have any cells left, or you have very little cells left, then you might be losing all your population on your negative plate.

If you do have cells in suspension, I would start to question your positive plate.  Are you making them yourself or are you buying them coated? Which rat strain are you using?  Perhaps you need to titer your antibody concentration on the plate.  There are really a whole lot of steps in immunopanning that can be optimized.


Why are VAL, MET and ALA substitutions commonly used for protein behaviour and function studies?

Substituting a single amino acid and checking the effect this has on a protein is a method to determine what this specific amino acid does in the protein. For example, substituting an amino acid that is part of the catalytic core would almost always make the protein non-functional.

Alanine scanning is a technique that methodically replaces amino acids in a protein with alanine. Alanine is probably the most neutral substitution, you essentially remove the amino acid sidechain. Glycine for example would be problematic because it is more flexible and can adopt secondary structurs other amino acids can't.

If you want to find out what a specific amino acid does, you don't want to introduce any other new effects when mutating it. Depending on what exactly you're looking at, substituting other amino acids is certainly possible. Some changes to alanine might be too drastic and you might want to substitute an amino acid that is more similar to the original one. This all depends on what exactly you are looking at.


How are whole Haplotypes for Sequencing isolated?

If your aim is to isolate the two haplotypes (chromosomes) maybe you should try flow cytometry. You might read this paper http://www.ncbi.nlm.nih.gov/pubmed/21181533 by Dolezel which is leader in the field, at least for plant chromosomes. I'm pretty sure that the applicability of the method depends on the physical properties of the chromosomes you have (especially their size distribution).


What kind of spider is in this pic I took?

It's a Spiny Orb Weaver, aka Spiny-backed Orb Weaver. Orb Weavers are a very large genus of spiders; this one is a Gasteracantha cancriformis. The spikes can be black or red; there are many colors as well.


Wasp nest destroyed by some big wasp like insect: is that the queen?

The wasp is a Ropalidia marginata

The assailant is a greater banded hornet .
There is a video of one attacking a wasp nest here: https://www.youtube.com/watch?v=BPLRpEkNv8Q

Hornets can attack wasp and bee hives and decimate them. In europe there is an invasive species of hornet from japan which is twice as big as the european hornet.

The abdomen of the hornet is about half of the size of the animal. The abdomen of the queen Ropalidia marginata is about 1/3rd of the animal, she has very similar markings as the hornet, it seems to have evolved as a copy cat that consumes larvae. http://drkrishi.com/ropalidia-social-wasp/

You must add location information to search for animals by region and find the species. I only knew it was india/pak because of the avatar.


Using Jpred to predict secondary structure

That depends on what your are trying to do. Apparently your query sequence is similar to that of proteins with a known 3D-structure. As it says on Jpred's result page (and help), in this case it might be worth looking at these homologues with experimentally determined structures for information on secondary structure. Most likely it will be more accurate than Jpred's prediction.

How much of the secondary structure you can use from the 3D-structure also depends on which parts align between your sequence and the ones in the PDB. You can show the alignments on Jpred's result page by clicking the button below 'Alignment of PDB hits to your sequence'. However, if you are going with annotations from the 3D-structure, it would probably be better to use SWISS-MODEL directly.

If you don't want to see those hits and just force a Jpred prediction, you can tick 'Skip searching PDB before prediction' in the advanced options before submitting your prediction. The hits are shown merely as a convenience, ignoring them doesn't change anything about Jpred's prediction.


Is post-transcriptional regulation of gene expression an epigenetic process?

miRNAs and other post-transcriptional regulators are very well "genetic". They are encoded by genetic elements, are expressed and are affected by mutations. Just because this mode of regulation was not well known previously, it should not be classified as an epigenetic mechanism while the traditional protein based transcription factors (TF) are not.

Epigenetics, as it is originally defined (the "formal definition") is about mechanisms that can perpetuate the state of a cell to its next generation. Inheritance of gene expression programmes is therefore epigenetic. Although the gene expression programmes themselves can be implemented via different factors including protein and RNA based regulators, they would not necessarily constitute the epigenetic mechanisms that lead to inheritance of this state.

rg255's point of view is that any mechanism that causes a variation in the functional aspects of the genome without altering the genome sequence itself, would be epigenetic. This is technically correct but in that case all gene expression regulators including TFs should constitute epigenetic mechanisms.

Now, the main issue is where to draw the line between gene regulation and epigenetics?

In my opinion, the epigenetic mechanisms are one of the ways to regulate the gene expression. Although histone modifications and DNA methylation regulate gene expression and also confer heritability to the gene expression programme, the heritability can be implemented without them as well.

You can imagine a cell as a vessel which runs a system of biochemical reactions. This system can have multiple steady states (for e.g. multiple fates of a stem cell). To perpetuate a state, the new cell just needs to have the right initial conditions. This can be proved mathematically too. Such a system can be implemented via the traditional transcription factors as well. So what is epigenetic?

IMHO epigenetic was a loose term to denote something that people were not fully aware of, at that time. Anything that was not directly mediated by transcription factors was termed as epigenetic, including long distance regulators, non-coding RNA etc.

BOTTOMLINE

I would not classify non-coding RNAs as "epigenetic" for the very reason that they are encoded by genes and have more or less a direct effect on the target genes, just like TFs (which are apparently not epigenetic). As for the papers, there were many papers that used to assign these under epigenetic mechanisms, but that is IMO just too vaguely arbitrary. (Ironically, I happened to come across miRNAs and lncRNAs while I was doing a summer project on epigenetics and was reading relevant papers.)

What should be considered epigenetic would be a subject of another debate.