text:
What is the difference between Founder effect and Bottleneck effect?
Both are examples of genetic drift in which there is a change in the allele frequency when the population size becomes small.
What is the difference between the two?

Genetic drift is an EVOLUTIONARY PROCESS
Source: Wikipedia > Genetic Drift

Genetic drift (also known as allelic drift or the Sewall Wright effect2 after biologist Sewall Wright) is the change in the frequency of a gene variant (allele) in a population due to random sampling of organisms.

I think that these terms are used without a perfect semantic. An obvious case
A population bottleneck is a DEMOGRAPHIC EVENT
Source: Wikipedia > Population Bottleneck

A population bottleneck [..] is a sharp reduction in the size of a population due to environmental events (such as earthquakes, floods, fires, disease, or droughts) or human activities (such as genocide)

A founder effect is an EVOLUTIONARY PROCESS THAT RESULTS FROM A FOUNDING EVENT
Source: Provine 2004

The “founder” principle, as introduced by Mayr in 1942, was an auxiliary mechanism (less important than random drift) for producing reduced variability in an isolated population started by a few individuals or even by a single fertilized female. Mayr saw a distinction between the “founder” effect and random drift. The founders were not a random sampling of the main population, but, once isolated as a small population, would undergo random drift

In other words, a founder effect does NOT refer to the ppulation bottleneck causing by a founding event. A founder effect refers to the increase in genetic drift (loss of genetic diversity is expected) caused by a founding event. A founding event is the event by which few individuals (eventually only one) quit a large population to settle down a new small populations. As such, a founder event is a type of population bottleneck. I think that one could further extrapolate the term "founder effect" to any increase in genetic drift caused by a population bottleneck.
Do all authors use the same definitions?
I have not done a good literature review to answer this question but I would not be surprised if different authors, use slightly different definitions.
For example, some authors (e.g. Peery at al 2012, Raymond and O'Brien 1993, Duarte et al. 1992) have used the term "genetic bottleneck" instead of "population bottleneck". Wikipedia indicates that these two terms are synonyms. However I could not find any paper that formally define the term genetic bottleneck but all the papers using this term seem to be specifically interested into the genetic signature of population bottleneck. As such, the semantic relationship between population bottleneck and genetic bottleneck could be the same as between founding event and founder effect. Genetic bottleneck could be defined as the increase of genetic drift that is caused by population bottleneck.

Founder effect and population bottleneck are not necessarily examples of genetic drift. The common thing in all three is a resulting reduction in genetic diversity.
Genetic drift is a change in allele frequencies due to process of random sampling to form a finite offspring population from parent population. Ultimately some alleles gets fixed and genetic diversity decreases. However decreasing population size or small population size is not a necessity for drift to happen.
Population bottleneck and founder effect are similar population genetic events that are happening in different ecological context. Both results in decreased population size (for the population of interest) and decreased genetic diversity. But the exact ecological context determines how much of the decrease in diversity is due to drift and how much of that is due to selection.
Population bottleneck refers to an event, where a population is intact but only small portion of the individuals are able to pass their alleles to future generations. This may be associated with strong natural selection, if possessors of a trait are more fit under the condition that causes the bottleneck (if the condition is a draught, better water preserving traits would be favoured for example). Or the process may be associated with drift if individuals don't have traits that would provide them an advantage (if the condition is a volcano eruption, survivors would be random with respect to their alleles).
Founder effect refers to an event when a small subpopulation migrates to an isolated region. Isolation may be due to distance or some geographic agent.
Here, again the individuals to form the subpopulation may be random with respect to their alleles, or there may be a selection towards traits that provide some advantage in migration for example.
I think that a population bottleneck is more likely to be associated with strong negative selection and a founder effect is more likely to be associated with genetic drift.
However these concepts are not about relative importance of drift and selection. (I emphasized my answer on drift/selection because the question considers them as examples of drift.) These both concepts are about the loss of genetic diversity due to decreased population size, and for that matter, I would say they are the same thing in different ecological context.
Edit:
You can refer population genetics text books by Hamilton (Hamilton, 2009) or Gillespie (Gillespie, 2004) for example. You can find related discussion on sections about effective population size.
There you can see that neither population (or genetic) bottleneck nor founder effect are treated as “examples” of genetic drift or defined as events caused by genetic drift.
Let me explicitly state my point.
Population (or genetic) bottleneck and founder effect are events (driven by some ecological factor) of sharp but temporarily decrease in population size. Due to this decrease, genetic diversity is decreased and stays low even after population size increase again. This is an example of the case, where effective population size and census population size are very different.
Now comes the part creating the confusion:
Both of these effects enhance genetic drift. But they are not caused by genetic drift. Genetic drift can be effective also when population size does not decrease.
Also they are not necessarily examples of genetic drift, meaning that, there may cases where the ecological factor causing the mentioned effects does not affect individuals within population randomly with respect to some of their traits. In that case some loci will lose its allelic diversity due to selection. But here again it is not selection that causes the mentioned effect, the cause is some ecological factor. The ecological factor causing the bottleneck may or may not cause selection. Population expansion/shrinkage is a dimension, selection/drift is another dimension.
Gillespie, J. H. (2004). Population Genetics: A Concise Guide (2nd ed.). Johns Hopkins University Press.
Hamilton, M. (2009). Population Genetics (1st ed.). Wiley-Blackwell.

So the bottleneck effect can happen due to a strong selection pressure too?? Interesting! Could you add in a reference?

Even though the reference I mention is not the most scientific one, it clearly differentiated selection and drift

You mentioned that the two effects are not examples of drift.Is it because natural selection has a role in bottleneck effect? My Text says they are.. So can you give a reference?

Ha! school mein thats how they taught.. Maybe this is correct. Lets wait for the reference :)

To emphasize again, genetic bottleneck or founder effect are caused by (are due to) neither genetic drift nor natural selection.

The Founder and Bottleneck effect are both examples of Genetic drift as you mention. The end result is pretty much the same, i.e., a reduction in the genetic diversity of the population. The main difference is the cause underlying the drift.
Say there's a very colorful ethnically rich and all population. As in, having whites, blacks, Chinese, Indians and all.. Suddenly two blacks are for some reason disappointed and migrate to an uninhabited island. The future generations would definitely be black. So the initial population which had all these Indians Chinese and all was definitely genetically more diverse than this population of only blacks. So this Founder effect lead to a reduction in genetic diversity (compared to the original population) - due to the fact that only the blacks chose to colonise.
On the other hand, the Bottleneck effect happens when there's a natural calamity. Say, there's an earthquake in the area where our rainbow population resides. All houses have collapsed and people have died except for the house in which the blacks lived. This happened just as a matter of chance. This is very important since it should be differentiated from natural selection. If the blacks survived because they were too smart to run out of the house during the quake unlike the others or if they were too strong to hold the roof even as it was collapsing, a component of selection would be introduced since they (or their progeny) would be genetically predisposed to do a similar thing if the earth shakes sometime later.
But here in genetic drift which is another explanation for evolution, different from natural selection, its just a matter of chance.
So all in all, Founder and Bottleneck effects are different in their cause not the effect which happens to be the same.
There are some real examples which I would urge you to refer online.
Here's a reference similar, simple and crisp
http://science.opposingviews.com/comparison-bottleneck-effect-founder-effect-5188.html

Do you have any citations/sources to support your answer?

Yup. Just adding them in :)

I wouldn't write my answer if I saw yours before I post. But I am not sure if these effects are defined such that they refer only to cases where selection is not taking place.

Thanks for the reference, but the confusion arises: does selection have any 'role' on bottleneck effect or is it just a matter of chance? That is where the difference is in the two answers.


text:
Can cell division be stopped during prophase?
There are three known checkpoints which ensure proper division of the cell.

These are: the G1 checkpoint, also known as the restriction or start checkpoint or (Major Checkpoint); the G2/M checkpoint; and the metaphase checkpoint, also known as the spindle checkpoint.

I wanted to know that is there any way cell division can be stopped during prophase(once it has started).

source: Stack Exchange

license: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/3.0/

sort: votes


text:
What was the first gene(s) found to code for ncRNA
I could not find a source that states what the first gene found to code for ncRNA was. Someone told me, however, it was a gene that coded for either rRNA or tRNA. To this point in time I have had no other confirmation that this is true.

The first eukaryotic genes cloned were the 18S and 28S rRNA genes from the frog X. laevis as described here: http://www.genome.gov/25520303
And yes, they are definitely ncRNA genes.

I don't think you answered my question. I asked what the first gene or genes found to code for ncRNA were. Not the first genes cloned.

I don't think you read my answer; the first eukaryotic genes cloned actually were ncRNA genes.

all_licenses: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/3.0/


text:
Why is sport healthy if high metabolism decreases life expectancy?
There is something that appears as a contradiction to a newbie like me.

Sport is widely recognized as healthy and recommended by medical experts.

Metabolic rate is inversely correlated with lifespan.

To me, it makes sense that the more energy passes through your system, the more it gets damaged.
Sport goes against this desirable frugality, as it makes you consume more energy, which requires increasing your energy input to compensate (you eat more food). Also, growing a greater muscular mass increases the amount of energy needed for moving, for maintaining body temperature at rest; having more cells mean larger amounts of oxygen are needed, more DNA-copying errors are made, more free radicals are produced, etc.
At the same time, I've heard that sport is good because it lowers heartbeat at rest, it makes special immune-system agents enter cells more effectively, and other things...
I'm obviously missing something, but I'm just having a hard time understanding how something that increases the entropy of your system can be beneficial for your health in the long run.

I think you're drawing extremely broad and simplistic strokes to draw conclusion number 2 and ignoring certain implications by the reasoning in your largest paragraph For example, large animals have to produce more cells to get to the size they are yet tend to have longer lifespans. Also, your use of the term entropy seems to be a misapplication of the term. The reasoning 'To me, it makes sense that the more energy passes through your system, the more it gets damaged" is also incomplete because it ignores how if too little energy passes through your system, you promptly die.

@DKNguyen Although it is also worth pointing out that larger animals generally have specific anti-cancer adaptations that help them in that regard.
By comparison, you can't just blow up a regular animal, and expect it to fare quite as well. Larger dogs are at greater risk of cancer compared to smaller ones, for example.

having more cells mean larger amounts of oxygen are needed

Two things here. First is that that a large muscle usually comes from the individual cells becoming larger, not so much that there are more of them.
Second, is that exercise means that your body uses the oxygen more efficiently if it's fit (it's unclear whether this refers to it having a higher concentration of oxygen, or whether it is due to changes in metabolic processes).

more DNA-copying errors are made, more free radicals are produced, etc.

Although it's worth pointing that these happen anyway. Exercise doesn't really change much on that front, unless there's an injury.
Your body has protective mechanisms against these forms of damage, since they happen as part of regular metabolism. They don't just gradually accumulate throughout your lifetime.

I'm obviously missing something, but I'm just having a hard time understanding how something that increases the entropy of your system can be beneficial for your health in the long run.

It helps stave off other health conditions that would take you out first.
System entropy increasing isn't really that big of an issue anyway, since humans aren't a closed system. A warm bath also increases entropy (from heat), but certainly doesn't cause health issues, and hypothermia tends to be quite bad for you, even if it does the opposite.

license: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/4.0/


text:
Fruit Fly Hybrids
I have a food waste bin in which I put fruit scraps. The fruits come from all over the world, mainly Europe though.
I'm in the UK.
I assume the fruit Fly eggs are already in the fruit, in which case they originate in the same country as the fruit.
Are they all the same species? Am I creating Hybrids in my waste bin?

@Remi.b your second statement is incorrect. While D.melanogaster is a genetic model organism, it is only one of hundreds of species in the genus Drosophila. Also your first statement is incomplete. It is the ability to produce fertile offspring that usually classified a species, as hybrids such as ligers or mules are, for the most part, sterile, though there is active controversy to the validity of the reproductive argument for speciation as well,

Probably they are not all the same species, as there are many fruit fly species in the UK and all over the world. It is also probable that you are not creating hybrids, as fruit flies have quite specific mating behaviours that change rapidly sometimes even between strains.
Anyway, you are assuming that the eggs come with the fruit, but that's hardly believable unless you already see larvae when you ate the fruit. Drosophila melanogaster has one of the fastest developments (22h since egg fertilisation, although it can vary with temperature). So, if we take for granted a week span since they were taken out of the country until you bought them (which is very optimistic) you would already see larvae in the fruit. In short, the flies that are thriving in your fruit bin are common flies from the UK.
In case you want to know, the most common flies in the UK according to this article is Drosophila subobscura and obscura, which are pretty similar in development to D. melanogaster.

Excellent thanks very much for your reply. Im amazed that the fruit flies are local species - there are so many in such a short time whenever fruit is left lying around. I guess you've answered that too!


text:
Do mutations come in pairs?
As DNA is organized in a double-helix, a change in one strand must be accompanied by a corresponding change in the other strand. Let's say an original sequence CATCAT mutates into CATGAT. The other strand must change from GTAGTA to GTACTA - or, more precisely, as the other strand is read in the opposite direction, from ATGATG to ATCATG, possibly in a completely different gene. Has the effect of this pairing of mutations been explored mathematically?

Welcome to Biology! What exactly do you mean with Has the effect [...] been explored mathematically? Mathematically implies you wish a theoretical analysis of the occurrence of mutated base pairs? Or do you want to know if indeed mutations are characterized by changed base pairs?

If a mutation has to be accompanied by a second mutation, only loosely related (by an accident of being on a strand exactly opposite the first mutation), it should affect a frequency of survival, or of a beneficial change. We should not consider a single mutation in isolation.

Note that DNA repair mechanisms catch odd bases. A mutation is per definition a change in base pair. Further, only 1 strand serves a basis for transcription. The other one does not. This question is quite unclear to me and I vote for closure until refined.

If a mutation occurs in one strand and is missed by repair mechanisms, it will be copied to the new complementary strand during replication. You'll end up with one double helix with the mutation and one without. That said, I also don't understand your question.

@AliceD This is a technicality, but rarely you will find two genes that overlap on complementary strands and so both strands serve as a transcription template at that locus.

@canadianer - we might have to take this to chat, but I always thought the complementary DNA strand was the template and the other coding strand just acts as a backbone. So the coding DNA strand carries the code, but does not act as the template during mRNA transcription. If that makes sense...

@AliceD You're absolutely correct, I was just referring to cases of overlapping genes, like this. So both strands at that locus act as a template for transcription. One of my previous professors claimed that essentially every part of the genome is transcribed at some point, which seems to be at least partially validated by papers like this.

While the question of mathematics makes no sense. The intention of the question is clear. What would happen if the DNA repair like nucleotide excision repair has failed to repair the DNA? I think the answer is that the mutated base will be replicated among with its new base-pair.


text:
Do ginko trees grow well in Eastern North America (specifically Ontario,Canada)
I have recently noticed many Ginko (Ginko biloba) saplings planted in my Eastern Ontario city (in Canada), and they are looking a bit sickly, with yellow leaves.
They seem to be a popular choice in new developments, but I didn't think they grew well in the region (which is in Hardiness Zone 5).  Could the yellowing leaves just be poor management, or are the poorly suited to the region?
Wikipedia tells me that although these trees are native to China, they have been cultivated in North America for 200 years.

It took a little bit of digging (no pun intended), but this seemed to be a resource provided by Tree Ontario/Forest Ontario. http://www.seedlingnursery.com/resources/item/21-choosing-the-right-tree . Your best bet is always to ask a nursery or parks service in your local area. They usually have professionals with a great deal of knowledge on local conditions and the best choices based on weather, growing season, soil conditions, drainage, and indigenous parasites.

I have now spotted several ginko trees that appear to be mature and healthy; and established in residential/park area rather than in an arboretum or educational/research facility.  I conclude that they must be able to survive here.

As a side note, ginkos are excellent at removing air pollutants and living in toxic environments. It has a wide range of temperature tolerance and is very hardy. It sounds like yours have a possible magnesium deficiency which is common in gingkos, or the plant might be over or under watered. A very helpful link all about the gingko, its habitats, and usefulness

@AMR I actually disagree (to...some degree). Though there are usually no other options than to speak to a nursery or parks service, these sources are often not accurate. Even experienced nursery employees are not always going to get it right. I have experienced this multiple times at nurseries in various US states. Further, while I worked as an environmental educator within the park system, I saw misinformation be presented on numerous occasions. I don't mean to discredit your comment at all, but rather just point out that those sources come with their own risk of inaccuracy.


text:
Is it possible for a human being to live on a different day cycle?
I am considering writing a story where a character lives on a 30-hour day cycle.
That is, this person sleeps for 9.5 hours and is awake for 20.5 hours.  Then this person repeats the schedule.
I'd like to know from those of you whom are more knowledgeable about biology and physiology whether this is possible.

Yes, it is certainly possible. Whether or not it's healthy is a different question!

Any chance this is motivated by https://xkcd.com/320/ ?

Ha.  Nice find.  I hadn't seen that one.

all_licenses: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/4.0/


text:
Does GABA help or hinder anxiety?
This article
https://www.prnewswire.com/news-releases/smartphone-addiction-creates-imbalance-in-brain-300558945.html
Says both

The researchers performed MRS exams on the addicted youth prior to and following behavioral therapy and a single MRS study on the control patients to measure levels of gamma aminobutyric acid, or GABA, a neurotransmitter in the brain that inhibits or slows down brain signals

And

Having too much GABA can result in a number of side effects, including drowsiness and anxiety.

If GABA slows down brain signals, wouldn't that reduce anxiety? And if you have too much GABA, wouldn't that reduce anxiety also, not increase it?
DreamWater, a 5 Hour Energy counterpart, has GABA, 5-HTP, and melatonin, which promote sleep. It wouldn't make sense that GABA increases anxiety to promote sleep.

I wouldn't bother trying to interpret press-release science that doesn't even contain a citation or link to the original research. However, I will try to address some of the other content of your question.
It is indeed true that many anti-anxiety medications, particularly the benzodiazepines, act primarily to increase GABAergic activity by potentiating GABA receptors.
However, it is also true that GABAergic activity is not some monolithic attribute of the brain. Different brain regions have different functions, and some brain regions are associated with behavioral or neurochemical inhibition of other regions. An increase in GABA activity in a brain region that tends to inhibit others could theoretically be anxiety-inducing rather than anxiety-calming, even if the overall effect of increasing GABAergic effects is anxiety-calming. Looking at GABA levels is also only one part of the picture, and it fails to tell you a) how often GABAergic cells are releasing neurotransmitter, and b) how sensitive post-synaptic cells are to GABA. The overall effect of GABA is going to be roughly proportional to (GABA concentration) * (how often GABA is released) * (sensitivity to GABA).
An extreme example of why this is important are the delirium tremens that occur if a severe alcoholic suddenly stops drinking alcohol. Alcohol also potentiates GABA receptors, and with long term use the brain compensates by reducing GABA production. However, this adaptation is fairly slow, and alcoholics who quickly stop drinking can have motor symptoms and seizures as a result (some other neurotransmitter systems are involved as well, though, so this is a bit of a simplification). The opposite could be true as well: it's possible that increased GABA concentrations could be because not enough GABA is being released or because postsynaptic receptors have been downregulated. It is quite possible that you could have more GABA but less GABAergic inhibition. This study didn't test levels of inhibition, just levels of GABA.
Lastly, eating GABA to make you sleep is a bit silly and is far removed from brain levels of GABA or inhibition in the brain - I am not aware of any study that shows that eating GABA makes you sleepy; companies can freely put it in a drink and make abstract claims that suggest it aids sleep, but that doesn't mean it actually does, a beverage like that isn't regulated the same way that pharmaceutical drugs are.
In summary: be very wary of attributing a complex behavioral outcome to one particular element in the brain.

I know, I know, I' not supposed to do this, but this is such a good answer, I have to. Thanks! I'd upvote more if I could.

@anongoodnurse Agreed. Just in case you are not aware of this feature, if you still feel like that in 2 days, you can add a bounty.


text:
Refer to Ran-GEF as a coenzyme?
Ran-GEF facilitates an exchange of GDP for GTP in the formation of Ran-GTP in the nucleus as part of gated import of proteins. Would I be correct in calling Ran-GEF or Ran-GAP coenzymes?

coenzymes are factors that bind to the enzyme so that enzyme can show its activity. In this case conversion between Ran GTP and Ran GDP are caused by hydrolysis and phosphorylation reactions. Ran GEF and Ran GAP are not binding to Ran GTP or Ran GDP.

Cofactor perhaps?


text:
Methods for phage separation
How can a mixture of unspecified phages be separated (into singular phage strains)? I.e, what are the main methods?
My research shows it can be done using CsCl centrifugation and affinity chromatography, but both seem laborious, the latter also requiring the phages to be already available in pure form.

Look up "phage purification." Basically, you add your phage solution on top of your host bacteria on a plate and incubate. As the bacteria grow, you should see plaques (clear areas in the bacterial lawn). Each plaque should be the progeny of one original phage, although sometimes two or more phages could be very close to each other on the plate and it will look like one plaque. So you have to take a sample of a plaque, re-plate on another lawn of host bacterium, and repeat this process for several rounds until the plaque morphology seems to indicate that there is only one type of phage present. Here is a good resource for phage protocols: http://phagesdb.org/workflow/

Thanks, excellent link! Any other known way other than manual extraction from plate? Is it perhaps easier to isolate infected bacteria during lytic cycle (or perhaps during induced lysogenic cycle)?


text:
Obvious phenotypic inheritance markers in humans
In humans, are there phenotypic markers that prove parenthood? For example, if mother and father have <trait characteristic> (let's say, specific ear lobe form), then any child will have <trait characteristic>. If the child does not exhibit that characteristic, than either father or mother or both are only alleged, not actual.

Interesting question. Eye and hair colour (two "obvious" ones), to my knowledge, can be different from either parent due to the dominant/recessive nature of alleles. I would be interested to know of any "true" , as you put it

I've previously heard claims that a cleft chin is such a trait, but this seems to be a myth: udel.edu/~mcdonald/mythcleftchin.html. The trait supposedly has a strong heritable component though. Blood type could sometimes be a strong indication of parenthood (e.g. an O father should normally not produce AB offspring), but not always.

Not that I know of.

No. To determine the genetic relationship (if any) between two individuals requires molecular evidence based on genetic markers in their somatic DNA. Using phenotypes could be misleading because a genetic marker (an allele) can have variable penetrance (the percentage of carriers who actually have the phenotype) and variable expressivity (the severity or degree of the phenotype). Some of this variability can come from gene-environment interactions, and some could come from epigenetic effects (ie non-heritable).


text:
Determination of animal and vegetal pole
How do we determine which is an animal pole and which is a vegetal pole in a zygote?
My professor said that the zygote can be divided into two hemispheres, vegetal and animal poles. The hemisphere with yolk is the vegetal pole and the one without is the animal pole.
But I couldn't find an authentic book that I have access to saying anything about the polarity of the zygote. However I stumbled upon this paragraph from Objective Zoology and I'm not sure if it's right.

Zygote or egg of frog is spherical and measures about 1.6mm in diameter. The upper half of the zygote is black (pigmented) and contains nucleus and cytoplasm. The lower part of the zygote is white or light yellow in colour and contains the reserve food in the form of yolk. The upper pigmented part is called the animal pole and the lower part vegetal pole.

"Vegetal" and "animal" poles are archaic terms. As pointed out by Elaine, the polarity axis is defined by ectoderm and mesoderm. There are other kinds of polarity as well. You can read more about it, for different organisms, from Developmental Biology by Gilbert.

To resolve this question here's something I found in Developmental Biology by Gilbert $-$

The amount and distribution of yolk determines where cleavage can occur and the relative size of the blastomeres. When one pole of the egg is relatively yolk-free, the cellular divisions occur there at a faster rate than at the opposite pole. The yolk-rich pole is referred to as the vegetal pole; the yolk concentration in the animal pole is relatively low.

Do you have any reason to doubt this statement? The wikipedia article says pretty much the same thing. When reading the folllowing, don't forget that one can talk about such poles at different stages of development.

The animal pole consists of small cells that divide rapidly, in contrast with the vegetal pole below it. In some cases, the animal pole is thought to differentiate into the later embryo itself, forming the three primary germ layers and participating in gastrulation. [..] The vegetal pole contains large yolky cells that divide very slowly, in contrast with the animal pole above it. [..] The animal hemisphere is dark brown, and the vegetal hemisphere is only weakly pigmented.

and it comes with the following picture

You know, wiki article talks about polarity of an embryo and not a zygote. I haven't encountered any authentic book saying that they are the same, so is my question.

Molecular Biology of the Cell, 6th ed, p. 1156 http://i.imgur.com/ZNTT53Oh.jpg After sperm entry and rotation of symmetrization, the zygote has an animal and vegetal pole. It also says elsewhere that the name animal and vegetal are due to old beliefs, and roughly correspond to what will be inside (endoderm and mesoderm) and outside (ectoderm) respectively, after gastrulation. The v-a polarity is established by gradient of stocked mRNA and vitellin. Mecanisms vary extensively between phylum.


text:
Null hypothesis of Chi-square test for independence
My understanding of null and alternate hypotheses :
The null hypothesis is a hypothesis whose plausibility is being tested by a test statstic and is denoted by H0. It assumes that observed data follows a standard scientific theory (and the variation/deviation is due to
chance).1 Alternative hypothesis is a hypothesis that contradicts null hypothesis. It assumes that the variation in the observed data is real, not due to chance.It is usually the hypothesis that a scientist is trying to establish.
(Source: Biostatistics by Lee & Forthofer & Statistics by David Freedman)
This1 holds good for Goodness-of-fit test but not chi-square test for independence as there is no theoretical data to compare the observed data with.
So my question is in a given problem of chi-square test for independence how to determine what the null should be?

When what the scientist is trying to establish is mentioned:
Q : In a study of effectiveness of an antipsychotic drug, patients are treated with the drug and were compared to those receiving a placebo. 698 of 1068 patients were released after taking the placebo while 639 out of 2,127 relapsed after taking the antipsychotic drug.
Test the prediction that the antipsychotic drug is significantly more effective in preventing relapse than placebo.
A:  Ho : Edrug = Eplacebo  Ha : Edrug > Eplacebo
(Reasoning: Ha is the hypothesis the scientist is trying to establish and so Ho should be something that opposes the Ha that he has to disprove.)
When what the scientist is trying to establish is not mentioned (at least not clearly):
Q: Among 60 males and 50 females, 25 males and 20 females were with attached ear lobes. Statistically prove whether or not the attached pinna has any relation with sex.
Not sure what the scientist wants to establish so unable to assume a null.

What is an attached earlobe? Earlobes are always associated with the pinna right?

@AliceD It's a commonly used human trait for biology/genetics classes that was thought to follow Mendelian genetics (attached being recessive). It refers to whether the earlobe is attached to the face along its length or whether there is a portion that hangs down below the lowest attachment point of the pinna. Its better known now that it actually isn't a simple Mendelian trait but it still shows up in textbooks as an example.

@SanjuktaGhosh I don't understand what your point of confusion is. There is a theoretical null when you use chi-square test for independence. Specifically, the null is that the variables are independent. So, in a population that has both males and females and attached and unattached ear lobes you expect the proportion of males with attached ear lobes to be the same as the proportion of females with attached ear lobes.

This question looks more appropriate for CrossValidated than Biology to me. The questions are framed as biological questions but this is really a question about statistics -- nothing fundamentally "biological" about them.

@BenBolker I agree. There was a post on this matter on Meta, and in fact according to it should still stay on Bio.

@BryanKrause Is it the same for homogeneity? P.S. Thank you. Given the time constraint I'm forced to do surface reading.

@SanjuktaGhosh Yes. Maybe part of the confusion comes from the somewhat poor naming of the tests. In truth, a "test for independence" is really testing for non-independence: that is, you reject the null hypothesis if the samples are not independent. Same thing with tests for homogeneity, normality, etc: in all cases, you are rejecting the null if the data are not homogeneous, not normal, etc.

@BryanKrause Here(1st problem, a one-tailed test) the Ho: Pdrug_released>=Pplacebo_released  Ha: Pdrug_released<Pplacebo_released ? My understanding

The Chi-squared test of independence is, as the name suggests, a test of the independence of two outcomes.
Two outcomes are defined as independent if the joint probability of A and B is equal to the product of the probability of A and the probability of B. Or in standard notation, A and B are independent if:
P(A ∩ B) = P(A) * P(B)

from which it follows that:
P(A | B) = P(A)

So in your drug example, there is a probability that a person in the study is given the drug, denoted P(drug), and a probability that a person in the study is released, denoted P(released). The probability of being released is independent of the drug if:
P(drug ∩ released) = P(drug) * P(released)

Release rates can be higher for individuals given the drug, or they can be lower for individuals given the drug, and in either case, release rates would not be independent of drug. So Ha is not
P(released | drug) > P(released)

rather, it is
P(released | drug) ≠ P(released)

In your second example, there is a probability that a person is female, denoted P(female), and a probability that a person has attached earlobes, denoted P(attached). These events are independent if:
P(attached ∩ female) = P(attached) * P(female)

and Ha can be stated as:
P(attached ∩ female) ≠ P(attached) * P(female)

or equivalently:
P(attached | female) ≠ P(attached)

Not sure if you missed this. The question in the 1st problem essentially asks to test the hypothesis that the Effectiveness is more. 'Test the prediction that the antipsychotic drug is significantly more effective in preventing relapse than placebo.'

That's the reason I pointed out that a significant result on a chi-squared test of independence is not sufficient to conclude that the drug is more effective than the placebo, only that the effectiveness of the drug and the placebo are not equal. Researchers will often get a statistically-significant 'not equal', and then look at the contrast and interpret it as 'greater than' or 'less than' based on the observed direction, but that is not what the test formally tests.

Could you tell me a bit more about 'Researchers will often get a statistically-significant 'not equal', and then look at the contrast and interpret it as 'greater than' or 'less than' based on the observed direction'?

Some detailed examples are available here: https://math.la.asu.edu/~elajack/Fall14/STP231/Lecture_ch10.pdf.
Essentially, since the test only formally tests 'is response independent of treatment', a researcher who gets a significant result might then look at their data to infer directionality. e.g., a researcher has 100 trials of treatment A, and 100 trials of B. The success-rate is 22/100 for A, and 80/100 for B, so they have a significant result: success-rate depends on treatment. Since 80 > 22, they might further conclude that success is higher for B than for A, with caveats at the link.

It might help if you first ask yourself what the independence test is for in plain English, you are testing to see if one categorical variable has
an influence on another. In other words, if a person (or whatever)with attribute X (for example, is bald) are they significantly more/less likely to have attribute Y (like being allergic to penicillin) than someone who does not have attribute X.
So in your example the alternative hypothesis would be (in words): the ratio of males with attached ear lobes compared to males without is equal to the ratio of females with attached ear lobes compared to those without.


text:
What is proton leak?
I have some idea about it. Is it the wasteful back flow of H+ ions through uncoupling proteins during aerobic respiration?
Thank you

Short answer: yes.  It also includes unspecific leakage without the aid of uncoupling proteins, since the mitochondrial inner membrane is not entirely impermeable to protons. For example, protons might fall backward through the proton-pumping complexes, through other solute carriers, or through other openings or irregularities in the membrane.


text:
Thermal ratchet model

Two models have been proposed to explain how ATP hydrolysis by mitochondrial hsp70 drives protein import.In the thermal ratchet model (Figure 12-28A), the emerging chain slides back and forth in the TIM23 translocation channel by thermal motion. Each time a sufficiently long portion of the chain is exposed in the matrix, an hsp70 molecule binds to it, preventing further backsliding and thereby making the movement directional. Thus, a hand-over-hand binding of multiple hsp70 proteins translocates the polypeptide chain into the matrix.

What does thermal motion mean here? I will be obliged if somebody could explain it.

It refers to the "random giggling" of molecules, which increases with increasing temperature (kinetic energy)

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.


text:
Why is tRNA called an adaptor molecule?
All I know about adaptor is that it is a device that converts attributes of one electrical device or system to those of an otherwise incompatible device or system.
Thank you.

When you make a statement like "why is tRNA called an adaptor molecule" you imply that it is some kind of a definition, which it is not. I can call tRNA using any of its attributes for example "carrier molecule" or "essential molecule" etc. You can check the meaning of "adaptor" in a biological context and see why it fits for tRNA. IMO, this question has a very narrow scope.

RNA and proteins are not electrical systems, but the idea of translating a signal between incompatible systems can be applied here.
Proteins are made through a process called Translation, where instructions stored in a piece of mRNA are used by the Ribosome to make protein.
tRNAs adapt the mRNA nucleotide sequence into a protein's peptide sequence by acting as an intermediary. A tRNA is charged by attaching an amino acid to one end, it then binds the ribosome mRNA complex at the position defined by a codon. A codon is a sequence of 3 nucleotides that codes for an amino acid.

You could think of the tRNA being like a SD Card to USB adaptor to get information from one device to another.

The term "adaptor" does not solely refer to electrical connections. In fact, any system that can establish a connection between two otherwise incompatible systems is called an "adaptor", as you can tell from the following list:

Electrical adaptor (mechanical incompatbibilty between power sockets/plugs)
AC/DC converter/adaptor (electrical incompatibility between electrical currents)
Pipe coupling adaptor (mechanical incompatibility between different pipe types)
Railway coupling adaptor (mechanical incompatibility between different railway couplings)
Computer periphery adaptor (mechanical and/or electrical incompatibility between different periphery links)

It is just natural that the term "adaptor" has been applied to concepts in biochemistry. You can't connect an amino acid to the RNA template directly. The two incompatible systems are thus RNA and the amino acid. The tRNA solves this problem. It is also not the only case where this term is used in biochemistry. Other examples include the signal transducing adaptor proteins or the vesicular transport adaptor proteins such as adaptin, that establishes a connection between the clathrin molecule and the cargo receptors during vesicle formation:

Francis Crick published the The adaptor hypothesis in a privately circulated article (now available online) “On Protein Synthesis” in 1958. He wrote:

“One would expect, therefore, that whatever went on to the template in a specific way did so by forming hydrogen bonds. It is therefore a natural hypothesis that the amino acid is carried to the template by an ‘adaptor’ molecule, and that the adaptor is the part which actually fits on to the RNA. In its simplest form one would require twenty adaptors, one for each amino acid.”

The word ‘adaptor’ has no intrinsic electrical connotation: the Oxford English Dictionary online gives the first definition and examples for this as:

Adapter (also adaptor)
A device for connecting pieces of equipment that cannot be connected directly.
‘the adapter converts the lens focal length to 5.5 mm’
‘a tube fixed to the cleaner by means of an adapter’

Crick clearly used the word in this sense.
At that time the idea was first mooted (reportedly 1955) it was not known that the adaptor molecule was transfer RNA (tRNA), although I have labelled it as such in the illustration I have prepared for the iPhone generation.

[Textual Explanation for the visually impaired
The illustration shows: 1. Chemical incompatibility: a three-pin electrical socket, labelled ‘mRNA triplet codon’, and the flat connection end of a USB cable, labelled ‘cognate amino acid’; 2. The Adaptor: an adaptor plug (labelled tRNA) with the three pins labelled ‘anticodon loop’, and the USB socket labelled ‘acceptor stem’; 3. Decoding: the USB lead connected to the adaptor plug, which has been inserted into the the three-pin socket.]

Not sure why this question resurfaced, but I thought that the accepted answer did not address the analogy, and the comment fails to acknowledge the historical use of this term. I have not included a diagram of tRNA showing the key features as it is assumed the reader has encountered this already. It is available in numerous pages on line, although best read about in a book.


text:
Is Plasmablast a precursor of Plasma cell?
I read it in Roitt's Essential Immunology.

Plasmablasts are precursor cells of short- and long-lived plasma cells and
  are generally described as a proliferating fraction of
  antibody-secreting cells, often found in the bloodstream
  emigrating to organs such as the bone marrow.

I coudn't find any authentic source repeating the same.
Is it true? Is this book authentic enough to follow?

I follow Kuby's for Immunology. I don't know about Riott's but Nature reviews would be the best for immunology.

@Polisetty Why do you think it is the best?

They have the latest stuff and amazing diagrams. Like the one below, even the subtle difference that the plasmablast has membrane bound Ig and plasma cells have secretory Ig has been made.

Antigen activated B-cells enter the germinal centre dark zone to form centroblasts which undergo somatic hypermutation. These then form centrocytes in the light zone. The various possibilities/routes the centrocyte can take are shown in the following figure. If the centrocyte undergoes a class switch recombination, it becomes a plasmablast.
A plasmablast is  defined as

Plasmablast
The B-cell lineage precursor of non-dividing plasma cells, which has the capacity to divide and that has migratory potential.

Once it matures as a plasma cell, it is out in the blood secreting soluble immunoglobulins or antibodies.
Another important definition to mentioned here would be of antibody secreting cells.

Antibody-secreting cells
A term that denotes both proliferating plasmablasts and non-proliferating plasma cells. The term is used when both cell types might be present

So, yes, they indeed are a precursor of plasma cells.
Also, it is necessary to clarify here that although both plasma cells and plasmablasts are antibody secreting cells, plasma cells lack membrane bound antibodies while plasmablasts retain them.

Finally, this is just the germinal centre response that is shown over here. Plasma cells and plasmablasts can also form in an extrafollicular response in extrafollicular sites in the spleen or in the medullary cords of the liver. That's another story altogether!
For a more detailed explanation, have a look at my other answer.
Image and reference: http://www.nature.com/nri/journal/v8/n1/full/nri2217.html

In your other answer you mentioned that they produce secreted IgM. This is also mentioned in Kuby. Is it an exception to what you said, they still producing membrane bound antibodies? Could you elaborate?

Thanks for pointing out. There's a small error. I'll update in a while :)

Both the answers have been updated @SanjuktaGhosh . Please have a look! Those plasmablasts are different extrafollicular ones

I appreciate that. However I regret not having access to the paid website, nature.com. :(

@SanjuktaGhosh Shouldn't you have access via your university?

@Ebbinghaus No I can't for my country is not enlisted there.


text:
How does one insert cas9 into animal cells?
How could cas9 be inserted into cells by researchers looking to edit a genome? I imagine for engineering bacterial systems you could just put in the cas9 coding region in an expression vector, but is that how it is also done for eukaryotic cells?

In short, yes. It's just under a mammalian promoter.

Any other notable adjustments that would need to be made?

I should clarify my earlier comment - it's under a promoter appropriate to the organism of interest. I work with mammalian cells, so I have "mammalian" stuck on my brain...

Right, I guess what I mean is, are there any amino acid substitutions or things like that from the bacterial cas9 sequence? Or does it generally fold nicely and work well just using the same ORF?

If you are talking about the nuclease functionality of Cas9, then you aren't adding the Cas9 gene into the genome, you are transfecting it on a plasmid. Usually there is either some form of permeabilization done to the cells, or the construct is inserted into a viral vector in order to transfect the cells in tissue culture. In multicellular animals, you would need to use the viral vector route as would need to be done with any gene therapy technique. Addgene is a nonprofit plasmid repository that has many of the CRISPR/Cas9 constructs that have been developed, and they have an excellent resource on their Website.
You wouldn't really want Cas9 active in the cell after it has done its job, as you increase your likelihood of off-target hits. The target shouldn't be in the genome anyway if the targeting event occurred, because you have either substituted in another piece of DNA or you have cut the DNA and the DNA repair mechanisms have introduced random nucleotides in order to repair the double stranded break.
Eukaryotic cells, for the most part, cannot maintain plasmids. The Cas9 gene will be transcribed along with the guide RNA. The resulting protein/RNA complex targets the sequence of interest and makes a cut. When the cell replicates the plasmid will likely be lost, but the daughter cells will inherit the genomic edit made by the targeting event.
If you are looking to insert a different gene at the target site, say a reporter into the genome, then you co-transfect a linear piece of DNA with homology arms around the cut site so that that donor will be inserted via homologous break repair.
One of the Cas9 constructs that has bene developed has disabled the nuclease activity, so that the Cas9/RNA complex ends up acting as a binding protein to suppress the activity you are looking to study without actually deleting or replacing the gene. In this case you may look to insert the Cas9 cassette, in which case you would use a recombination event as you would with any other type of incorporation into the genome.


text:
How does NHEJ cause indels?
I was reading up on CRISPR-cas9 and how it works and I am having trouble wrapping my head around how NHEJ to repair the DSB can cause indels to occur. Shouldn't the NHEJ just stick the two strands of DNA back together?
I believe I read somewhere that if other agents cause a DSB, such as certain chemicals, they can also damage / cleave nucleotides ; the damaged nucleotides would have to be removed and overhangs filled in by polymerase before the ligation, which could result in the errors that NHEJ is famous for.
Since cas9 does not cause such damage, how can it result in an indel?
Thanks.

This is a very interesting question and I had not thought about it before. I am reiterating the background of your question.
Background
Cas9 cuts the both the strands of the target DNA at the same location thereby making a blunt end cut. Usually sticky ends and microhomologous ends lead to indels because of exonucleolytic clipping and addition of extra bases by pol-µ. The question is how do indels arise during NHEJ mediated re-ligation of blunt ends.

NHEJ may not always lead to indels. Most blunt end cuts can be repaired without error. Some non blunt cuts are also perfectly re-ligated; the best example for this is VDJ recombination. The binding of Ku proteins is essential for a perfect re-ligation (canonical NHEJ/C-NHEJ).
However, there are some variant non-canonical pathways which include alternative-NHEJ, backup-NHEJ and microhomology mediated end joining (MMEJ) (Bétermier et al. 2014). Some studies also refer to all of these pathways as alternative-NHEJ or altNHEJ. These alternative pathways involve resection of nucleotides by exonucleases such as Mre11 to produce overhangs. Imperfect alignment of these overhangs prior to ligation leads to indels. Sometimes non-templated addition of nucleotide happens by the action of terminal deoxyribonuclotide transferase (TdT). Pol-µ can also add random nucleotides at the 3' end (Lieber 2010).
In general, the propensity of DSBR via alternative NHEJ pathways would be higher when Ku activity is lower. Even with a moderate Ku activity, there is always some likelihood of alternative pathways in action. Moreover, it is to be noted that sticky end ligations are more efficient than blunt end ligations.

 Reproduced from Bétermier et al. 2014


text:
What does it mean to clone a gene?
When I look up information related to the identification of disease genes, texts will often refer to the gene being "first cloned." What does "clone" mean in this context? Is it simply a synonym for discovery?

Wiki > molecular cloning might help you.

Possible duplicate of Are there multiple meanings to "clone" as in "clone a gene"?

@Christiaan I would rather close that question as a duplicate of this one. Chris' answer below is much more informative than the one in the linked question. And yes, it is perfectly "legal" to close older questions as dupes of newer ones - we do it on Stack Overflow all the time.

@MattDMo, agreed. I retracted my vote.

@Christiaan I just submitted my duplicate/close vote on the other one - feel free to join in :)

@MattDMo - done :)

Usually this is used in the context of gene analysis and further characterization. What is done, is that a piece of foreign DNA (either generated by digesting genomic DNA (from human or mouse for example) or by PCR amplification) digested at the end of the DNA to have matching open ends with a bacterial vector.
The DNA of interest is then ligated with the vector and can then be replicated in bacteria and isolated easily in great yields. This plasmid can then further analysed (by restriction analysis, sequencing etc.), the gene or its promoter manipulated and mutated, or in case of protein coding genes the gene overexpressed in the cells. All this is useful to analyze the function of unknown genes. See the figure for a schematic overview (from  Campbell AP Biology Book 8th edition, via this website):

Technically making a clone means that you generate multiple genetically identical daughter cells from one precursor.

I think that might be a figure from Campbell Biology. I know you put a link, but you may want to attribute it... as it is copyrighted material. Figure 20.2 in the Ninth Edition, page 397.

Oh, I didn't know that - I have adjusted the information.

I have seen Campbell so much in my life that I can pick their graphics out from 150 meters... :-) Same with Molecular Biology of the Cell and Janeway's Immunology. I can usually spot Nature's cell biology graphics as well. They are all very branded and distinctive.

That's right. I never used the Campbell very much...

So basically, the act of cloning the gene is more or less necessary to sequence and characterize it, thus when people say that a gene is first cloned, they're effectively referring to its full characterization?

In the lab it means that you identified and isolated a certain sequence.


text:
How are at home fermentations without starter cultures controlled?
When making fermented food at home, particularly those that are made without the addition of a starter culture, how does one ensure that the correct types of bacteria are carrying out the fermentation? Is this generally due to the properties of the food that is fermented?

can you give an example?

@MattDMo I think a traditional kimchi preparation is a good example. You've added no bacteria of your own except for what you obtain preparing the dish from its ingredients, and then you seal it and let it ferment at room temperature. I think in this case the acidity is what keeps the food pathogens from growing.

A "starter culture" is just a culture of preferred organisms. They probably differ little from local wild organisms. For example, a mixture of flour and water left to stand will (with luck) be colonized by yeast in the air that result in a serviceable yeast culture. Control of this process entails nothing more than observation over time. The environment and nature of the medium suffice to control the bugs that colonize it.
Both the Korean and Chinese versions of kimchee depend on moisture and acidity (and perhaps salinity) to inhibit unwanted molds/bacteria. The process is evidently self-controlled, but I recall seeing a film in which it was said that failure to add salt to the mixture could result in spoilage. Note that in the linked article, the author says cavalierly that one can scrape any mold that forms off the cabbage and continue with the fermentation because mold will not grow in liquid--patently false but probably "true enough" not to affect results.
Aspergillus oryzae colonizes rice, and rice with Aspergillus growth can be subjected to a secondary yeast fermentation that yields sake. The commercial production uses highly-prized starter cultures for both the mold and yeast, but it is possible to substitute other varieties of Aspergillus. For example, one can induce A. niger to grow on a citrus peel, and transfer the colony to rice. The sake that results is fine. But in this case one would want to rule out species of Aspergillus that are associated with toxins. For similar reasons some care is taken in sterilizing containers used to make wine.
In any of these processes there is a possibility of colonization by the wrong flora. The cultures are controlled by the content of the nutrients and climate and observed carefully. Over centuries there have probably been many unfortunate accidents involving unwanted organisms. There is nothing like violent illness to bring the integrity of a process into focus.
In short, the control of home fermentations lies in (1) statistical likelihood that the right organism is present in the environment; (2) the nature of the nutrient and climate; (3) the experience of the person(s) monitoring the process; and (4) luck.

I see. So essentially, conditions are created that likely favour the desired fermanting organisms, and spoilage, if it occurs, might be detected via smelling/looking off, etc.

That seems to generally be true, yes. It's a broad question but I think people acquire a lot of experience and can tell by smelling, tasting, looking. It's probably not infallible.


text:
What is the role of glucose in plasmid isolation?
My understanding is that glucose is used in the resuspension solution to prevent cells from bursting by maintaining the appropriate osmotic pressure. Why do we even bother doing this? The cells are going to be lysed with the addition of the SDS+NaOH, and we have EDTA in the resuspension solution to chelate Mg, thus inactivating nucleases.

This is a common misconception: in fact, in the alkaline lysis method for plasmid isolation, the glucose does not act as an osmotic stabiliser.

Glucose is present in the resuspension buffer at 50 mM. This concentration is not high enough for osmotic protection of spheroplasts formed by lysozyme treatment: from memory, 0.5 M glucose is typically used for stabilisation. So why is the glucose included?
The original method for plasmid preparation by alkaline lysis was published by Birnboim and Doly in 1979. They explain that the method relies upon achieving a pH of between 12.0 and 12.5 after addition of NaOH so that linear dsDNA (sheared chromosomal DNA) is denatured but closed-circular DNA (plasmid) is not. They state in the paper that:

further pH control is obtained by including glucose as a pH buffer.

According to this site glucose has a pKₐ of 12.3, which aligns nicely with the claim that it helps to keep the pH within the desired range.

This is an interesting bit of information (I really like the explanations of this mechanisms). Thanks for finding it.

@Chris I actually knew the answer, having been around when the Birnboim & Doly paper was published. This technique revolutionised what was possible with plasmid-based cloning at the time and I read it very carefully. At the time of writing (05-07-2017) the paper has 12,877 citations, but most people who use the technique (in kits) won't even know of its existence. Sic transit gloria mundi

The kit problem (as I usually call it with our students) is exactly the reason why I love the information bits. They help in trouble shooting and it is good to know how a method works anyway. Classical biochemistry with finding out exact mechanisms. I thought I would know the lysis method and the steps pretty well, but there is always something to learn. As it turns out, this paper is already in my literature database, I will re-read it sometime later.

Really great answer, but the second link is broken for me.

@canadianer fixed, I hope.


text:
Why is there a size limit on inserts that a plasmid can accept?
Throughout my undergraduate education I have been taught that plasmids can't carry very large inserts, but I have never been told why. Any insight would be greatly appreciated.

Transformation efficiency and the bacteria's ability to maintain the plasmid is reduced as the plasmid size gets larger. Bacterial (300kb) and Yeast (1000-3000kb) Artificial chromosomes can be pretty big, but it is more difficult to get them to work. Also most of what you are working with won't be that big. Inserts usually come from cDNA produced by PCR from mature mRNAs. Without introns, the majority of genes you will see won't be more than a few KB, so plasmids do not need to be that large.


text:
Why is western blotting used to confirm positive ELISA HIV tests?
I know that when an ELISA test indicates a positive result for HIV, a western blot is done to confirm. When I search online for the reason why, all that I see is that western blotting is "more specific" than ELISA. If they both use antibodies, shouldn't these procedures have similarly accurate results?
My only guess would be that the combination of Ab-Ag binding in addition to expecting your antigen to be a certain size is sort of a "double yes" and thus western blot results would possibly be taken as more accurate.

The only other thing I would add is most commonly used ELISA employ an indirect approach of detecting Antibodies in the serum against HIV antigen attached to the ELISA plate

One reason a Western blot is more specific than an ELISA - even one using the same set of antibodies - is background.
Antibody-linked colorimetric reactions aren't completely on/off. There's always some background reaction. The antibodies are a little bit cross reactive, or non-specifically sticky, or the colorimetric coupling enzymes aren't completely washed away, etc. There's a number of reasons you'll get a small amount of signal even in the abscence of the desired reagent.
So imagine how this non-specificity looks in an ELISA assay. You're only getting a single readout. Any non-specificity/background in the assay adds noise to your output level. Hopefully for any decent assay the noise is normally well below the threshold for crossing over into a "positive" readout, but if the noise is Normally distributed, there's always a small chance that the noise will exceed that threshold. (There's also those non-normal failure modes, where you mess up a wash step, or you get a bad tube of antibody, etc. Negative controls help, but only so much.)
In contrast, think about how non-specificity/background looks in a Western blot. If you get non-protein linked background, the presence of this background will be across the entirety of the blot. It's very obvious that something went wrong with the assay in those cases. Even if you get random spots and not overall background, the chance you'll get a random spot at just the correct molecular weight is very, very unlikely.
An additional benefit is that Western blots first separate the proteins prior to doing antibody-linked detection. This means if there's some protein in the sample which cross-reacts with the antibody, it's very likely it will migrate at a different molecular weight and not be counted as a false positive. (As opposed to ELISAs, where every signal counts toward the total.)
These effects mean that if you're looking for low false positives (as you would with an HIV test) a well-done Western blot is likely to give a more confident signal than a well-done ELISA would. (At the cost of time and effort.)
A final issue is that combining the two will give you more confidence in a positive result than either one alone, especially if you use different antibodies for each. The final confidence in a positive result is not just the confidence from the Western blot, but the confidence that both the Western and the ELISA report positive results, so there's a benefit of doing both, even if they were equally specific.


text:
How can the melting temperature of PCR primers be so far below the extension temperature?
Since our primer TM is lower than the extension temperature by design, (and often by a significant amount) shouldn't the primers be released from the template at extension temperatures?
Now, I have done PCR experiments and know they work. I just want to justify what's actually going on molecularly to be at peace. The only thing I can think of is that as the temperature ramps up to the extension temp, the polymerase has already begun adding nucleotides, thus raising the effective TM, preventing dissociation.

That will happen in some cases, but by the time you are ready to extend, the annealing step will have mean that primers formed double helix stretches with you template, and the polymerase will have bound to both, stabilising it, thus it isn't free to be released from the strand.

Note that 'melting temperature' is actually about POPULATIONS--it's generally the temperature at which 1/2 of all of the millions of copies of 2 things will be stuck together and 1/2 apart. Once stuck, not all primers will leap off the template simultaneously even at much higher temperature... As you indicate, there's nothing to stop the polymerase from starting its work (extending the primer) before the extension temperature.
And, as in another answer, polymerase can add stability to the pairing.


text:
Using Q solution with ready made MasterMix
I am exploring the possibility of using Q solution (5x) to get rid of non specific bands in PCR. I mostly use a MasterMix and not separate aliquots of dNTPs, Taq, buffer etc. In principle, adding Q solution to the reaction mixture when I use MM should be fine, but I just wanted to ask if someone has done so and had success (or failure?) before.


text:
BSA for trouble shooting failed PCRs?
I am using PCR to amplify a 1000bp region of the CytB mt gene. Some of the samples had not amplified and I wish to change the reaction components (or use additives like BSA) to attempt at getting amplification. The DNA was extracted from liver/muscle samples, so I am not expecting many inhibitors present, but I am eluting samples in AE  buffer.
I used the following combinations of PCR mixture but the reactions still failed (reaction volume 20ul)
Combination1 : MasterMix (10ul), Primer 1 (1ul), Primer 2 (1ul), BSA 2mg/ml (2ul), water (4ul), DNA (2ul)
Combination2 : MasterMix (10ul), Primer 1 (1ul), Primer 2 (1ul), BSA 2mg/ml (4ul), water (3ul), DNA (1ul) : Tried to increase BSA concentration and reduce DNA to decrease the volume of inhibitors (if any) going into the reaction mixture.
Someone who has previously used BSA to successfully troubleshoot PCRs, am I going about this in the right way ? If I need to take a different approach please help me out.
Thanks!

The albumin shouldn't be what your trouble shooting I don't think. It's something I've always kept constant if required by the enzyme. Since your inconsistency is across different samples and I assume you are creating a 'mastermix' containing the taq, ntp's, primer etc I would troubleshoot the purification steps. If that's not possible, there are other parameters of The pcr (especially annealing temp and primer concentration) that will have more apparent results. MtDNA usually requires a more stringent extraction protocol like using guanadine.

There are other samples from the same extraction batch that are working fine, so I'm assuming the problem is not with the extraction or with the PCR temperature settings ? And I use a mastermix, yes.


text:
How do I use BEAST for divergence time estimation in this case?
I am interested in using BEAST to find the divergence time estimates of species separations within a genus of frogs. The genus separated from a outgroup taxon I have chosen around 80 million years ago. That is all the information I have. Is it possible for me to do a divergence analysis using BEAST and set an informative prior based on the information I have ?

Doesn't BEAST require molecular sequences for it to do its work?


text:
Effect of 260/230 values on PCR
To what extent does the 260/230 value affect the efficiency of PCR reactions ? Do the constituents leading to a low 260/230 value (EDTA, guanidine salts and oligosaccharides) inhibit PCR ? I have been getting pretty low values of 260/230 (0.5-1.0) for DNA extracted using Charge Switch Protocol using Magnetic beads and the PCR has been failing. Also, is there any reason why I might be getting low 260/230 values in Charge Switch method ? (I was getting good values using Phenol-Chloroform extraction method)

I've never heard of the charge switch method, I've only ever used phenol chloroform extraction and isopropanol precipitation. If phenol chloroform worked for you before, why not go back to it?

EDTA would certainly inhibit PCR.

@user137 charge switch  is adjusting the ph to change the charge of the magnetic binding bead, change in charge allows washing and elution

I doubt it would matter. 1 ul template in a 50 ul reaction - you've diluted it 1:50. There's nothing left - and you could easily go down to 0.2 ul for 1:250. Also, your enzyme makes a big difference. I would email the enzyme manufacturer and ask them.

I use a 10ul or 20 ul reaction actually, still I get your point.

EDTA carbohydrates, phenol all have absorbance near 230 nm.  TRIzol reagent is a phenolic solution which absorbs in the UV both at 230 nm and ~270 nm
Guanidine HCL used for most DNA isolations will absorb at ~230 nm also
All these things will seriously inhibit your pcr, (and lower your ratio) especially the denaturing agents like guanadine.
All these DNA kits are different spins of a pretty similar process.
If you have a good steady centrifuge stick to the phenol & ethanol. Be certain you are drying your pellet well, but don't over dry it. I'll hear my pellet at 37 for 3-4 min to get rid of ethanol. Ethanol no good in pcr. If you hate doing ethanol precipitation (which I do) cause of invisible pellets, add a high grade bit of glycogen to your aqueous phase before the precipitation. This will make your pellet visible and is completely inert.
If your cleanup isn't the problem do an annealing  and Mg gradient and up your primer quantity.
May want to give me more info like the individual absorbance, pcr conditions, template size etc
Traditionally bead purifications gives really good purity, reason I use it for microarrays. But it's expensive and not necessary for pcr.
Charge switch is just changing the charge of the bead by changing the ph.

My PCR thermocycler conditions are as follows -
Initial Denaturation - 95/30s
[ Denaturation - 94/10s, Annealing - 57/60s, Extension - 65/9m30s ] x 35 cycles
Final Extension - 65/9m30s
I am trying to amplify a 10kb fragment from the mitochondrial genome, using NEB LongAmp Taq Polymerase.
I use 10ul reactions - 5ul LongAmp MasterMix , 0.4 + 0.4 Forward and Reverse Primers (final 0.5mM in reaction), 1ul BSA (5mg/ml BSA), 1ul Mg2+ (final 1mM in reaction) (optional), 1ul template (amount 40-80ng) and make up volume with RNAase free water.

If 260/230 is a problem, it should inhibit all PCR, but I am not getting results for long fragment amplification specifically. For around 1200 bp using Qiagen Hot Start Kit, I am getting results.

Your amplifying a pretty big fragment, your going to need close to perfect conditions. 1200bp and 10kb are pretty different in terms of pcr. Also I would check your protocol of how you are getting the mtDNA, you may need to enrich your template by doing a mito purification. Let me know if this makes sense

Also your extension times seem too long that pcr reaction must be taking half the day. What does NEB recommend for the extension?

@anurag mishra what's the melt temp of your primers?

Melting temperatures are around 60C-62C and NEB recommends 1min/kb of amplicon, hence the extension temperature. Yeah, it does take 7 hours!

So with that reaction time, your reaction really has to be crystal clean. Even trace amounts of contam would cause probs if exposed for that long. be sure your using the highest grade water, dntp's and desalted primers, That your template is fresh and without knowing the number of reactions you are running I would scale up the overall volume to increase your precision (even if that means a little waste).

Should be running multiple to compensate for fidelity and sequencing issues I would think anyways. Hard to say in this case the template gets so diluted it's rarely an issue in shorter reactions.  Check your dntp concentration it may need to be a lot higher for such a big frag. And as always double check your primer seq's good luck man


text:
DNA extraction from agarose gel
Because DNA is soluble in water, is it possible to extract PCR product by dissolving excised gel containing the DNA band of desired length in water, mashing the gel piece with a pestle, centrifuging it and then pipetting out the supernatant ?

If you mash it with a pestle, you'll likely break the DNA. You can use a low melting point gel and mild heating followed by centrifugation to extract DNA.

why not use this? (http://www.qiagen.com/products/catalog/sample-technologies/dna-sample-technologies/dna-cleanup/qiaquick-gel-extraction-kit) or a similar product? I have used it many many times and it can be done very easily!

@Bez - Well, that would cost around 1-2$ per sample, correct ?

@AnuragMishra indeed, however, there might be delivery charges but some times institutes bulk buy and sell it in their own stores and their prices are less and sometimes there are institute based purchase pages which sell the products at the discounted price. ps I'm not promoting any brand or products here and I just put it up as a point of reference.

Yes, this is possible (and I have done it uncounted number of times) - the method is called "Freeze and squeeze". What you basically do is to run the gel, cut out the band of interest (be careful with the UV light, it causes damage to your DNA and also sunburns, so wear appropriate shielding for your face), dissolve it in a buffer, then freeze it in liquid nitrogen (or the -70°C, doesn't matter) and centrifuge it at room temperature for 10 minutes. During this time the ice melts and the agarose pellets at the bottom. Take the supernatant, do a ethanol precipitation (with glycogen when you only have little DNA) and you have nice and clean DNA. No need for expensive kits and is done approximately in 2 hours.
You can find a very nice and detailed protocol here: "Elution of DNA from Agarose Gels" (The rest of the handbook is also very useful).

Yes, we used to call this method 'freeze-squeeze'.

Right, I added this. Useful method and always amazed our new masters students. "What, no columns?" :-)

But, how will Qiagen make money if people know how to do this?

I have just put a comment about the Qiagen product. I guess ease of use of kits has made me very dependent. Its always great to find alternative methods to use. Many thanks for the method Chris.

@Chris instead of ethidium bromide use for DNA detection with UV, you can use SafeView Nucleic Acid Stain by NBS Biologicals, which uses fluorescence excitation maxima at approx 290-320nm, emitting at 515nm so its non-UV detection and it is very sensitive in my experience and safe too!

@Bez: Excitation 290-320 means that it needs to be excited in UV-B (which can cause DNA damage). The only difference is the emission wavelength which is green not orange/red. And that they claim that they are safer than EtBr.

@MattDMo I don't think that companies like Qiagen are in danger of going bancrupt any time in the near future. :-) Besides that, I started to use this, when we had problems of isolating short DNA pieces from the gel with columns. Subsequently I dropped columns for this work.

Extracting from PA gels is more pain. In such cases electroelution is also an option

PA gels are not dissolving when they are subjected to heat. But we rarely use them.

Ethanol precipitation is a overnight step, right ?

@AnuragMishra No not necessarily. Have a look at this and this article here at stackexchange. They go into detail. In short: Overnight brings no advantage.

The centrifugation after keeping the buffer+gel slices at -70C should be at how many RPM ?

@AnuragMishra At maximum speed, 14.000 or whatever your benchtop centrifuge can do.

Sorry to dig up an old post, but how does this method compare to the qiagen spin columns? When using spin columns I always have terrible 260/230 (< 0.1), suggesting that significant agarose is carried over. My yield also seems poor, but this could be related to the amount of DNA used in the gel. I usually do gel purification on PCR products and get <500 ng of DNA, but I can't calculate a percent yield because I don't know how much PCR product was there. But other people in the lab use the columns to gel purify 100 ug of digested DNA and get 50% yield, but don't tell me the 260/230.

@user137 Ihave never compared this systematically with spin columns. And I have mostly used this technique with digested DNA which was subsequently cloned (so mostly digested PCR fragments or pieces of DNA), so I generally had not much material in the beginning. Recovery was ok and cloning with this purified DNA was also not a problem.


text:
Possible reasons for DNA getting stuck in well
I am in the process of Library preparation for Illumina MI-Seq using mtDNA. Using NEB Hotstart LongAmp Polymerase, I was able to obtain upto 10kb amplicons of mtDNA. However, when I switched to a different DNA sample, I am not noticing bands anymore in the gel. Strange thing is that, if I fragment these samples (Diagenode Bioruptor acoustic shearer) I get fragmented smears similar to the ones I expect from fragmenting a 10kb amplicon, even though these are stuck in the well.
Therefore, I dont know if a 10kb amplicon is stuck in the wells and is not migrating (given I see similar results for fragmentation for the samples that got stuck), or I am seeing in the wells what is genomic DNA and no amplification has occurred. Does the 260/230 value affect PCR amplification in any way? (I have a very high 260/230 value ~5 for the new DNA sample I am using)

What kind of gel (agarose, acryl amide) and which percentage do you use?

I use 1% agarose gel.

I don't know enough about your gels to know what's making your bands stick, but I've seen bands stay in the well when binding DNA to certain proteins and peptides, but I do it intentionally. Could something be bound to your DNA? Could the DNA in your wells be larger than you assume, but fragments under sonication to make small enough fragments?

I have put forward some possible solutions/trouble shootings below however @user137 raises very interesting points, which should be strongly considered! I have put a potential solution to potentially address this point including the DNA size issue.

May I ask how you solved this problem? I got same problem. 6kb stuck in wells but ladder runs ok.even 0.7% gel didn't help

There is one simple reason for that, your agarose gel is most likely too dense. Depending in the type of agarose, I would prepare a 0.5-0.6% gel at maximum.

Synbio gives this list for "standard" agarose, which fits pretty good with my experiences. If you use low melting agarose, this table looks a bit different, as the gel matrix is not a dense. The disadvantage is that the gel is much more prone to get damaged while handling.

I was getting the band at 10kb corresponding to 10kb molecular marker before. So I am not too sure if the density of the gel is the issue here. Also, even when the sample is stuck, the molecular marker ladder runs fine.

The problem is that this really depends on the agarose (and there is a plethora of different brands available). I would say that you are in the region where it gets difficult to get a good resolution. If your fragments are getting a bit bigger, than they will get stuck and you get something when fragmenting it. I would try a lower percentage gel and see, how this looks. Ideally with the same sample that made problems for you.

@caseyr547 This doesn't mean that I am always right.

Given that the OP knows the behaviour of markers in this gel it seems unlikely that this is the problem. Also although 1% agarose may not be suitable for achieving good resolution in the 10 kb range, this is very different from a fragment not entering the gel. I would try cleaning up the template further.

This is hard to judge without seeing a gel image. When the library preparation didn't went well, the DNA might be bigger in size.

I think this is a case where Occam's razor can be applied. The problem appeared with a new DNA sample, so that sample must be the prime suspect.

That's correct. I would run it on a lower percentage gel to see what happens. If it is then still not working, it is probably better to prepare fresh DNA and stop riding a dead horse.

First as you mentioned, which is I think the key thing, is that you need to clarify what DNA sample it is that you are observing in the gel. The best thing to do to ensure that you only have PCR generated DNA samples is that once your PCR is over, treat your DNA mixture with Dpn I enzyme, which cuts methylated DNA, which is essentially cellular DNA as they get methylated in cells and it should not effect you PCR generated DNA as it won't be methylated. Look at page 7 and page 12 of this document for further instructions on using Dpn I (http://www.chem.agilent.com/library/usermanuals/Public/210518.pdf). You can subsequently clean and isolate your PCR generated DNA using any kit such as QIAquick gel extraction kit (QAGEN) using essentially this protocol but you are not using any gels (http://sites.bio.indiana.edu/~chenlab/protocol_files/agarose_gel_extraction.pdf). You can use other kits but this always worked for me! (please note that I'm not promoting any kits here!!) Now if you have any amplicons from your PCR then you should be able to nanodrop it (using the 260/230 ratio as your guide) and get a reasonable value for your amplicon concentration.
Now if at this stage you still have your PCR DNA as indicated by nanodrop and perhaps a test digest still gives the expected bands then the problem is with the running process and highly likely your gel which was the first thing I suspected to be the case. If not then you do not have any amplicons and you are simply trying to run your original DNA.
Although I have never worked with very large cellular DNA molecules before (I work with plasmids), however I was trained to run very large DNA (in agarose gel) on very low voltage such as 10-50 V overnight in a cold room since large cellular DNA doesn't tend to run very well in gels although that mostly applies to chromosomal DNA but 10kb is still quite large. I have run plasmid DNA of 12 kb in gels before at higher voltages but low voltage is best and it avoids tearing and shearing the DNA!


text:
RNA seq: Should non-infected units be used for RNA-seq?
I am designing a RNA-seq experiment to study transcriptomic profile changes upon parasite exposure. Some papers I read, mostly extract RNA from the entire treatment group (organisms exposed to parasite), without (seemingly) bothering if all the organisms were indeed affected. Doesn't this practice bias results? Are there any studies on how this could affect results?

It will dilute your signal if you have a pool of samples +/- the parasite prepared as one library prep. It is always better to maximize the number sequencing libraries (usually cost prohibitive) and extract RNA from samples you are confident exemplify the condition of interest, and an equal number of control samples to maximize your statistical power.


text:
How do nicks in the DNA strand affect the success of Long Range PCR?
Long Range PCR using NEB Master Mix - Hot Start Taq was working fine for me (amplicon sizes of ~10kb) but stopped working all of a sudden. Is it possible that many freeze-thaw cycles on the DNA template introduces nicks into the DNA strand that hamper the polymerase activity ? In certain cases, I am getting no amplification and in some I am getting very weak amplification. Because the integrity of the DNA should be intact over the entire length in case of long range PCR, am I right in thinking nicks in DNA will be especially detrimental if we are trying to amplify large fragments ?
The concentration of the DNA templates I used was 90-140ng/ul in a 10ul PCR reaction, DNA eluted in TE buffer after a magnetic bead method of DNA extraction.

It is not unlikely that nicking is causing trouble. The question is if repeated freezing and thawing is causing the problem, there is an article which has been published in B.R.L. Focus in 1983 which disputes this effect. You can find the PDF with the article here, the article itself starts on page 10 in the PDF. On the other hand there is this paper ("Impact of long-term storage on stability of standard DNA for nucleic acid-based methods."), which shows that sstorage conditions do have an effect on the effciency of qPCR standards. Whatever causes nicking, it is a problem in (long) PCR. The following quote is from Sigma in their long PCR enzyme mix:

Template High quality and adequate length of the template are
  essential for reliable amplification of larger fragments. Extreme care
  must be taken in the preparation and handling of the DNA target for
  long PCR. Nicked or damaged DNA can serve as a potential priming site
  resulting in high background. Avoid freezing, or, alternatively,
  freeze only once to minimize damage. The condition of the target DNA
  is critical. Depurination during cycling is minimized by use of
  buffers with a pH greater than 9.0 at 25 °C.

This paper ("Effect of Highly Fragmented DNA on PCR") shows that induced nicks affect the efficiency of PCR (although they do a much shorter amplification than you). So to overcome this problem I would make small aliquots of fresh DNA preps and then see, if this helps.


text:
Does exercising increase the rate at which UV rays are absorbed and cause affects?
Does exercising while in sunlight affect the rate at which the human body absorbs and reacts to UV rays? As sunburn is the secretion of fluids, increased dilation of blood vessels and inflammation of skin would exercising (as it also increases blood vessel dilation for heat loss) accelerate the "sun-burn" process in skin?

reacts? probably. absorbs UV? no.

The only pathways in which I would see more sensitivity to UV light would be

If you were drenched in sweat and somehow the sweat focused the light/UV.   This is highly unlikely as I don't believe any studies have been done to show that a water based solution causes a more intense sunburn.

(Here is a further explanation by another poster on wetness and sunburn https://physics.stackexchange.com/questions/71263/why-does-wet-skin-sunburn-faster )

The increased dilation of blood vessels results in a more red thus darker and more absorbent skin tone leading to more of a sunburn

Both of these are highly unlikely of increasing the sunburn, so in my opinion exercising shouldn't increase UV absorption or make a sunburn worse. Hope that helps.


text:
How can a two genera share a latin name?
Recently, someone asked if species could share a latin name. In his answer, Remi.b said the latin name is unique to the species, and I think he is right.
However, this made me remember that I once stumbled upon a latin name that is shared between a genus of birds, and a genus of plants: oenanthe.
https://en.m.wikipedia.org/wiki/Wheatear
https://en.m.wikipedia.org/wiki/Water_dropwort
My question is: how come? Is it allowed for genera to share a name (not likely)? And why did Linnaeus, who named both, choose to do so? Error?

No, not an error. Plants, Animals and Bacteria each have their own independent nomenclatural codes. So while it is not advised, it is not forbidden by the code to have the same genera. It would be forbidden to have the same genus within one of the three codes. As a side issue, don't forget that the species ALWAYS includes the genus. Species are always binomens. The species epithet is the second word in the species binomen. Thus our species is Homo sapiens, not sapiens.

The different nomenclatural code is probably because they belong to different kingdoms in the Linnean taxonomy.


text:
Is it possible for two people to repopulate earth?
Today I was wondering this question whether it is possible for two people to repopulate Earth.
What if everyone else disappeared, except for two people (man & woman)? Can they repopulate? In that case, what are the odds against and in favour of that ever happening?

Possible?: of course. Probable?: hmm... . Would it generate a healthy human population?: No.

Possible duplicate of How many people are required to maintain genetic diversity?

You cannot predict that. They may or may not lead to an unhealthy population of descendants but surely they'll face the risk of extinction because of lack of diversity. Extinction is dependent on the environment and it may or may not be adverse to this population. Since there are so many unknowns here, the answers would most likely be based on guesses. Hence I am closing this question as opinion-based.

Well, mathematically it might be possible. But, I would rather worry about the low amount of genetic diversity that the growing population would have, which would make it extremely sensitive to external stresses.
To repopulate earth from such a small pool of genetic material, one would need an organism that is much more sturdy as compared to humans in extreme conditions, multiplies fast and is much more prone to mutations that actually convert to a phenotypic significance so as to increase the genetic diversity of the total population.

Update (after receiving many comments)
Before you read answer, note that this is hypothetical question and I am trying to answer it by creating hypothetical situations with some reasonable numbers
Short answer: Yes, hypothetically it is possible for two people to repopulate earth.
Quoting from Popsci website:

Maybe, but they'd be very busy

Hypothetically it is possible but practically it has very less probability. Let us calculate probability of repopulating earth to it's 10% population.

Let us assume one female can give birth to 50 children throughout her reproductive life spam and assume there is no death/disease/natural crises and other factors which might restrict this 'ideal' growth.
To get 'maximum' number of births in next generation, assume she gives birth to 1 male child and 49 female children. Probability of having that is $\left(\frac{1}{2}\right)^{50}$.
Now again assume all 49 females gives birth to again same ratio of children (too much work for male :P). Probability of happening this will be $\left(\frac{1}{2}\right)^{50+49}$.
If you go by same logic, you need at least 5 generations to make 10% of earth's population. That gives probability of $\left(\frac{1}{2}\right)^{50+49+49+49+49+49} = \left(\frac{1}{2}\right)^{295}  = 1.57 \times 10^{-89}$

It is much lower than probability of we finding electron in nucleus :P
Now if you want to consider little bit realistic model where each couple will have two more couples and so on, It will need at least $2^{30}$ generations to repopulate to 10% of today's population. Considering average lifespan of 60 years, it will take $2^{30}\times60 = 6.4 \times 10^{10}$ years to do that. It is higher than entire biological evolution time scale.
Point of these calculations are to show that, hypothetically repopulating earth with one couple is possible but very less probable.

5 generations for all those people seems very reasonable space of time and isn't the real issue of "is it possible". I think the answer here needs to approach genetic bottlenecking, survival scores of individuals over extended periods of time, and optimum conditions required for repopulation.

I doubt in this hypothetical situation genetic bottlenecking will matter and I don't know how will you implement survival score here.

But if we are here in these numbers within some few million years when homo appeared - how would you explain that? And that probability calculation for 1:49 male:female is not to the point here.

Down vote ? It is hypothetical question. I have also mentioned in answer that "Point of these calculations are to show that, hypothetically repopulating earth with one couple is possible but very less probable"

The probability statements in this answer are not very precise. Let X(t) be the population t years after the human population is reduced to two people. What are the probabilities you are interested in? Pr[X(1000) > 0]? Or maybe the limit of that probability as t goes to infinity? Or the probability of reaching some threshold population in a certain amount of time?

When you write "Let us calculate probability of repopulating earth to it's 10% population", for example...  in what time frame?

@Adrain Well, in first calculations I don't consider time period because biologically that kind of dynamics is not possible. In second calculation I have mentioned biologically plausible dynamics with time period.

Correct me if I am wrong: in your first example, haven't you merely calculated the likelihood of 1 male descendent per female (while having 49 female offspring per female) over 5 generations? Yes, this chance is minuscule, but what has this to do with repopulation - apart from that this might be the "ideal" growth curve? Also - from a genetic point of view, a 50:50 male/female ratio would be a lot healthier.


text:
Seeing strange things in light
I have noticed that if you look directly at tubelight (I have only white ones in my home) and close your eyes little, you see little random shaped colourless things sometimes with slow speed and sometimes with falling down.
They are shaped like circles with something in between and in chains of random length.
My father has also seen them. So, I am sure I am not imagining things. What the hell are they?

Almost everyone has these and they are harmless in small numbers. If you have a sudden increase in the number of floaters then that is symptomatic of potentially serious problems and you should consult an eyecare professional.

It's been suggested that this is off topic here and belongs on [biology.SE].

Those are floaters. These are objects floating in the vitreous humour that fills the eyeball. They typically look like:

Thanks. Up until now, I knew that it were white blood cells.. :)

@Awesome: Have you tried Googling to find out? Given that this is a physics site I doubt there are many experts on eye health hereabouts.

@Awesome I can recall being fascinated by this when I was still in my cot - probably about 3 years old. I recall quivverring my eyes to make them move around! I'm now 50 and everything sight wise still works well!

In https://en.wikipedia.org/wiki/Blue_field_entoptic_phenomenon this phenomenon is explained with white blood cells in your retina.