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IOF World Ranking system
Posted on | January 3, 2014 | Category: News
The IOF is in the process of commissioning a new service provider for the IOF World Rankings in all disciplines. 6prog, who has maintained the World Rankings for 13 years, has indicated that they do not feel able to continue to run the rankings in 2014. President Brian Porteous comments: "We regret but respect their decision on this matter and I want to thank them for their years of loyal service to our sport."
The details of the new service will be announced shortly. Until that, information needed for ski orienteering to complete the season will run normally on the 6prog site.
The World Ranking system for foot orienteering has been changed for 2014. The principles were discussed at the Presidents' Conference, and after hearing the national federations' views, the Council approved the new system in October 2013. The basis for the changes for 2014 were to recognise the importance of our most prestigious events as a major contributor to the World Ranking Scheme, to ensure all nations who conduct a World Ranking Events will have participants contributing to World Ranking Scheme and to improve the value of our Regional Championships as a contributor to the World Ranking Scheme.
To find out more about the new system for Orienteering World Ranking, please read the news articles from November 2013:
Orienteering World Ranking System will change in 2014
No more zero points at World Ranking Events
The World Ranking top 50 as of 31.12.2013 can be found here:|| Print page ||


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Disclosures Tribunal: 'No evidence of any conspiracy or plan to harm Maurice McCabe'
13/02/2018 - 13:38:00Back to Charleton Tribunal Ireland Home
Update 1.38pm: "A series of most unfortunate events" led to the circulation of a false rape allegation against garda whistleblower Sgt Maurice McCabe in 2013, lawyers for the Garda Commissioner have have told the Disclosures tribunal, writes Gerard Cunningham.
The tribunal is hearing closing submissions from legal teams in relation to its first module, where evidence was heard in July 2017.
That module set out to investigate the creation, distribution and use by TUSLA (formerly HSE) of a file containing false allegations of sexual abuse against Sgt McCabe sent to Gardaí in 2013, and whether these false allegations and the file were knowingly used by senior garda officers to discredit Sgt McCabe.
A "cut and paste error" led to a more serious allegation being added to a report from Miss D after she sought counselling in 2013. Miss D had initially made a complaint against Sgt McCabe in 2006.
After a garda investigation the DPP decided that the allegation did not constitute an offence.
Tribunal chairman Mr Justice Peter Charleton said this morning that a GSOC review of the original garda investigation had concluded it was a thorough inquiry. He said the investigation, headed by Garda Superintendent (then Inspector) Noel Cunningham "would be a very good example for teaching purposes of how to conduct an investigation of this kind."
Micheál P O'Higgins SC, counsel for the Garda Commissioner, said that HSE officials had acknowledged that the errors which were made in 2013 were theirs.
"This was a series of most unfortunate errors," Mr O'Higgins said. He said the errors were not garda errors, and garda officers took reasonable steps to correct the errors once discovered.
"Perhaps chaos theory was ruling, rather than any conspiracy," Mr O'Higgins said.
The barrister said there was "no evidence the false accusations were knowingly used by members of An Garda Síochána to discredit Sgt McCabe in any way."
Mr O'Higgins said that in appointing Assistant Commissioner Derek Byrne and Chief Superintendent Terry McGinn to look into policing issues in the Cavan Monaghan division, the force made an appropriate and reasonable attempt to address Sgt McCabe's complaints.
Paul Anthony McDermott SC, on behalf of Tusla, said there was no evidence of any conspiracy or deliberate plan to cause harm to Sgt McCabe.
Mr McDermott said the agency wanted to apologise to those affected by the sequence of errors, and to apologise again in public to Sgt McCabe and his family. He said the agency was revising how it dealt with historic allegations.
Michael Cush SC, for the HSE, said there was "a terrible error", and when it was discovered HSE employees took steps to recover copies of the report which had been sent out, but unknown to them, a report has also been forwarded to An Garda Síochána.
Earlier: D family asked why sexual allegations against Maurice McCabe were being raised years later
The Chairman of the Disclosures Tribunal has asked counsel for the D family was there anything behind reviving allegations of sexual assault against Maurice McCabe in 2014.
The DPP had ruled in 2007 that even if there weren't issues of credibility with Ms D's account of events, what had occurred did not amount to sexual assault or even assault.
Peter Charleton asked counsel for the D family why the allegations were being raised again so many years later, including speaking to journalist Paul Williams.
Counsel for Ms D said she was concerned that Sgt McCabe was being portrayed in a glorified manner by certain sections of the media, and she thought there were different perspectives on matters.
- Digital Desk

subset: commoncrawl


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2 Good Lo took a leap of faith in quitting his job last year to hone in even more on music. The rapper didn't waste any time in the studio after making this decision. "In March, I dropped 'The Good Life: Season 1', and shot two videos for that. Then, in August, I dropped "The Good Life: Free Admission", and then dropped three videos for that," said the artist. Months later, 2 Good Lo is back to bringing the heat with "The Good Life: Season 2", which he released just last week. In this method of releasing music, the rapper has seen a steady increase in his following with each project he drops. His social media is a true testament of this, as the followers are flooding in at an astounding rate.
Fans can find his newest mixtape on all digital music streaming platforms. His single, "Waves" featuring Rico Nasty, has been keeping his fans steadily buzzing, as it became an instant hit. The entire visual aspect down to the cover art for his latest project is completely unreal. "In the center of 'The Good Life: Season 2' album art, it's me wearing a crown, because that's what I'm chasing. On the left side, you have me in the white robe, and on the other side, I'm wearing a red robe. So, my conscious is playing a role in this whole mixtape as well. I'm speaking through myself in the project." He plans to continue this process of accompanying each musical project with a visual to match.


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LA Furniture – a 180 degree turn on Phone Call Tracking
I am about to tell you a quick a case study/story of a complete 180 degree change in stance about Phone Call Tracking by one of my clients, LA Furniture.
The initial research
LA Furniture, nationwide online retailer of modern furniture and with Four furniture stores across Los Angeles, has been using CallTrackingMetrics since March 2013. It was I, Varun Tandon from Rockin Technology Web Marketing, who conducted a vendor research at the time. I researched through top 5 vendors for Phone Call Tracking. I narrowed my research down to testing two vendors. Over a span of 2 months, I rigorously tested out ResponseTap and CallTrackingMetrics side by side on two separate client's websites. Needless to say, CallTrackingMetrics won out of the two.
At LA Furniture, we ported numbers from other two existing phone vendors. If we were unable to port numbers, we slowly replaced them after a few months.
Initial use of CallTrackingMetrics (CTM)
I was in a happy place. Using New Phone Calls and Call Sale as an important indicator of paid & organic campaigns evaluation, I produced better results. It helped me evaluate advertising channels that were not working. Given that more than 30% of LA Furniture's sales were over the phone, we ended or extended advertising contracts based on additional data we gathered from CTM.
After almost 2 years of using CTM, LA Furniture started questioning whether the Return on Investment made sense on CTM's cost. As a marketing professional, I clearly saw it essential to my work as I used it each month for Adwords keywords evaluation, but it was never directly used by anyone at LA Furniture.
For over two months, we had conversations around CTM's worth. After many conversations, we got to a point where we started looking at all the communications system at LA Furniture.
After testing out LA Furniture's existing phone exchange menu system, we learnt that there were gaping holes where customers/callers were sent into a loop of menus at certain times during the day and during off hours.
Due to the missing control and knowledge about customer service phone system, we learnt that they could not effectively help certain customers.
The 180 degree turn
Instead of talking about CTM's worth and to get rid of it to reduce costs, we scheduled a screenshare call with Bob from CTM. He answered all our questions and showed us the use of Schedule times, Agent Queues and Voice Menus.
LA Furniture has since:
- LA Furniture been testing and setting up all Agents with Browser phones to answer calls.
- Teaching customer service reps on how to take notes within CTM so all Agents can help any customer.
- LA Furniture has come up with innovative hold music by talking about their ongoing sales while customers hold.
- All Agents can now check voicemails and follow up on customer calls, something that was almost impossible to do with the old system.
- Thus far, CTM has completely handled complicated rules around voice menus and scheduled timings.
Important note: LA Furniture's old phone system still exists, but it's bypassed completely. This was possible only because all of the advertised phone numbers are through CTM.
LA Furniture is very optimistic about huge improvements to their customer service and furniture sales department.
Till now, Rockin Technology had used CTM to provide an accurate ROI number for various campaigns for LA Furniture. Now, CTM is part of client's everyday work flow. LA Furniture's monthly payment to CTM remains the same while they clearly see it's worth as they use it each and every day.
Takeaway: As a marketing professional, you sometimes need to go outside of your role and help the client understand all the features that can be used to help their bottom line. Look at your client's phone system to seek opportunities. it may seem like a daunting task as it usually involves multiple people across the company, but it's completely worth it.


text: Item number KUAV7Features hyaluronic acid and gives a subtle dewy smooth finish that lasts all day Why You'll Love It Our air-whipped liquid foundation doesn't sink into pores but floats onto skin for perfect coverage and 100% poreless look. Glides. feels, looks like satin perfection. Ideal for normal skin. Oil-free, fragrance-free, Dermatologist-tested, allergy-tested, non-comedogenic - won't clog pores, safe for sensitive skinMaybe She's Born With It. Maybe It's Maybelline Goes well with Dream Touch Lumi Baby Skin Instant Pore EraserFor Best Results Apply smoothly and evenly to your face and blend with your fingertips. For Adult use only


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Today is also the day when House Speaker Nancy Pelosi and Senate Leader Harry Reid will hold an enrollment ceremony to send the recently-passed Iraq bill to President Bush, which he will veto tonight and before tomorrow's meeting with Congressional leaders on a new version of the bill. A few presidential scholars talked of the vote:
William Howell, a war powers expert and associate professor at the University of Chicago, said whatever the historical significance of last week's vote, Democrats have gained considerable traction in opposing a wartime president because of the war's unpopularity.
"It establishes this marker so that not now, but six months from now ... Democrats can have the momentum to (override) a presidential veto" if the war is still going badly, Howell said. "Just because it doesn't pass doesn't mean it's not of consequence."
Zelizer agreed, adding that an anti-war vote is no easy task when U.S. troops are fighting abroad.
"It's harder to extricate yourself from a big war, not just strategically but politically," Zelizer said. "It's that first vote that's sometimes the hardest."
Indeed, the article goes on to say the passage of the legislation in many ways surpasses congressional efforts to end the Vietnam War. Congress went years before it was able to agree on legislation significantly challenging presidential war policy, holding some 94 roll call votes on the war between 1966 and 1972.
One can only hope Powell is right that a few months down the road enough Republicans will defect to override President Bush (unfortunately, these are months many of our troops in the field don't have). In the meantime, his rejection of this bill now shows only that he is still as out of touch with reality of the Iraq war as when he declared "Mission Accomplished" four years ago. Any chance he'll pull out the flight suit again for the veto?


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Last Sunday I celebrated by 25th wedding anniversary with my lovely wife Claire. It is unbelievable to realize that I have now been married a quarter of a century! Where has time gone?
Getting married was a mind boggling experience for me as I had given up by the time I met her at age 36. But it was love and magic at first sight…pretty much…as I was immediately captivated by her boundless energy, enthusiasm, intelligence, and beauty. We were soul mates almost immediately and I had longed for one of "them there thangs" all my life!
A college psychology professor of mine once described marriage as an opportunity for a young man to "be redeemed by the love of a good woman." That redemption started immediately and will continue to the end of my life as redemption is always an ongoing process, a process which is best conveyed by the poet Wendell Berry in the poem I will conclude with shortly. One dimension of this redemptive process is just the simple structure provided by the commitment of marriage, two separate individuals with separate agendas deigning to live together under one roof. "commingling" their lives.
And when a man and woman begin to live together under one roof, that is when the sparks begin to fly! And they have certainly flown and it is a wonder at times that the entire house did not burn down! For marriage is work and if it is done "right" the work will challenge one to the core. Someone once noted that we marry a fantasy and when we get "in the saddle" together, the fantasy begins to dissipate and we discover the reality of the other person. AND, in the process we make a parallel discovery of the "reality" of who we are ourselves.
This "self" discovery is the most important gift that marriage has given me. As the work of marriage unfolded, I began to realize the true meaning of the biblical description of man and woman together as being "one flesh." I began to see how Claire was truly my complement, embodying so many things that I wanted but also so many things I did not want! And, of course, being a professional…and compulsive…"care giver", I wanted to "fix her" which I came to discover meant that I wanted, and even demanded, that she be just like me. Well, I soon learned that hell would freeze over before that would happen!
It probably took twenty years for me to learn just exactly what relationship is, to see…and feel…that Claire and I were "one flesh." I had to learn to make space for her in my life, to make room for her in my heart, and that entailed that I had to understand that I had not been doing so in the first place! And, even more so, it meant that I had not been doing so with anyone! I had known about the notion of "otherness" for a long time but suddenly the experience, or "feeling," of "otherness" was on the table and that was, and still is, disconcerting to say the least. Suddenly I was face to face with the subtle narcissism that had shrouded my life since early childhood. And slowly, and even shyly, I began to peer out of that shroud and to discover not just Claire but the whole of God's beautiful creation. I had understood Karl Jung's notion of "withdrawing our projections" but now I began to "feel" it also.
Here are two poems that so beautifully capture the mysterious work of marriage:
by Wendell Berry
How hard it is for me, who live
in the excitement of women
and have the desire for them
in my mouth like salt. Yet
you have taken me and quieted me.
You have been such light to me
that other women have been
your shadows. You come near me
with the nearness of sleep.
And yet I am not quiet.
It is to be broken. It is to be
torn open. It is not to be
reached and come to rest in
ever. I turn against you,
I break from you, I turn to you.
We hurt, and are hurt,
and have each other for healing.
It is healing. It is never whole.
And the second poem conveys the redemptive dimension of living together within the confines of marital commitment:
by Edgar Simmons
The stutterers hesitation
Is a procrastinate crackle,
Redress to hot force,
Flight from ancient flame.
The bow, the handclasp, the sign of the cross
Say, "She-sh-sheathe the savage sword!"
If there is greatness in sacrifice
Lay on me the blue stigmata of saints;
Let me not fly to kill in unthought.
Prufrock has been maligned.
And Hamlet should have waived revenge,
Walked with Ophelia domestic corridors
Absorbing the tick, the bothersome twitch.
Let me stutter with the non-objective painters
Let my stars cool to bare lighted civilities.


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It was a year ago that Lucy was first seen by a geneticists. She had been hospitalized at Primary Children's and we begin working with an incredible team of doctors who have trained us and loved us. We have learned so much. Lucy has been amazing through it all and so have her big sisters. I know many prayers have been offered in our behalf, I am so grateful for the love and support we have received, your prayers have sustained us.
Her geneticists was very diligent, exploring so many different ideas, then this spring, he was moved into a different area in his department, and Lucy was assigned a new geneticists. Lucy had started to make some great progress, she passed her swallow study, she started to get more mobile... so I wondered if we even needed to go meet the new geneticists. We did go, and the minute our new doctor saw Lucy, he said she reminded him of another baby he knew 11 years ago, and he thought she had Sotos Syndrome. The test results came back a couple of weeks ago confirming what he had suspected. So many things make sense, I understand better so many of the struggles that Lucy has been working on. It feels like we have met so many of the doctors and therapists right at the right times we needed to.


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West Aldin
The Six
The Six refers to the six gods of the pantheon. Although rarely interacting with mortals directly, they do speak to a chosen few. The Six are also responsible for the creation of many of the beings in the world.
Altanis the Just
Altanis created the versatile and honorable humans. In order to keep peace and justice, one must be able to see both sides of a coin – to be able to approach a problem from any and every angle.
Ignis the Bold
Ignis created the brave and hard-headed dwarves. Master of the flame, Ignis passed on his metalworking abilities to the dwarves, who became the finest craftsmen in the land.
Sylvanas the Patient
Sylvanas created the grounded and curious halflings. Knowing that only the fittest survive, Sylvanas made the halflings deceptively harmless, but with the cunning and speed needed to ensure their survival.
Aerathis the Wise
Aerathis created the strong and clever dragonborn. Modeled after the cunning dragon, the dragonborn use both their wits and their brawn to leave their mark in history.
El'Sulta the Hunter
El'Sulta created the swift and thoughtful elves. As the head of the pantheon, and the hunter of The Six-Headed Wolf, El'Sulta gave the elves prowess in archery, and control over the powers of the sun – light and fire.
N'zoth the Beast
N'zoth is believed to have created the beastmen of the world, such as the orcs and the goblins. A cruel and deceiving god, N'zoth is the shadow in the corner of your eye, or the being behind you, that always disappears when you turn around.


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On Show – Newlands Village (R5,3 million)
This immaculate 4 bedroomed home is set in the heart of Newlands Village with mountain views and has a garage as well as off-street parking for 2 cars. The home has a gracious style and the elegant and sunny living areas lead to a patio garden with mountain views. Beautiful finishes.
On Show this Sunday, 13 July 2014 from 14:00 to 17:00
Follow boards from Kildare Road or Newlands Avenue into Governors Lane
Property Sales Advisor: Lynne Boothe
Mobile: +27 (0)82 455 6467
Email: firstname.lastname@example.com
View more details of this property ENV71851


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The BARD1 gene encodes for the BRCA1-associated RING domain (BARD1) protein. Germ line and somatic mutations in BARD1 are found in sporadic breast, ovarian and uterine cancers. There is a plethora of single nucleotide polymorphisms (SNPs) which may or may not be involved in the onset of female cancers. Hence, before planning a larger population study, it is advisable to sort out the possible functional SNPs. To accomplish this goal, data available in the dbSNP database and different computer programs can be used. To the best of our knowledge, until now there has been no such study on record for the BARD1 gene. Therefore, this study was undertaken to find the functional nsSNPs in BARD1.
2.85% of all SNPs in the dbSNP database were present in the coding regions. SIFT predicted 11 out of 50 nsSNPs as not tolerable and PolyPhen assessed 27 out of 50 nsSNPs as damaging. FastSNP revealed that the rs58253676 SNP in the 3′ UTR may have splicing regulator and enhancer functions. In the 5′ UTR, rs17489363 and rs17426219 may alter the transcriptional binding site. The intronic region SNP rs67822872 may have a medium-high risk level. The protein structures 1JM7, 3C5R and 2NTE were predicted by PDBSum and shared 100% similarity with the BARD1 amino acid sequence. Among the predicted nsSNPs, rs4986841, rs111367604, rs13389423 and rs139785364 were identified as deleterious and damaging by the SIFT and PolyPhen programs. Additionally, I-Mutant showed a decrease in stability for these nsSNPs upon mutation. Finally, the ExPASy-PROSIT program revealed that the predicted deleterious mutations are contained in the ankyrin ring and BRCT domains.
Using the available bioinformatics tools and the data present in the dbSNP database, the four nsSNPs, rs4986841, rs111367604, rs13389423 and rs139785364, were identified as deleterious, reducing the protein stability of BARD1. Hence, these SNPs can be used for the larger population-based studies of female cancers.
Citation: Alshatwi AA, Hasan TN, Syed NA, Shafi G, Grace BL (2012) Identification of Functional SNPs in BARD1 Gene and In Silico Analysis of Damaging SNPs: Based on Data Procured from dbSNP Database. PLoS ONE 7(10): e43939. https://doi.org/10.1371/journal.pone.0043939
Editor: Amanda Ewart Toland, Ohio State University Medical Center, United States of America
Received: April 29, 2012; Accepted: July 27, 2012; Published: October 9, 2012
Copyright: © Alshatwi et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: This study was financially supported by the Research Centre, Deanship of Research, College of Food and Agricultural Sciences, King Saud University, Riyadh, Saudi Arabia. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
A single-nucleotide polymorphism (SNP) is the most common type of genetic mutation. There are several publically available databases for SNPs, such as dbSNP, GWAS Central, SwissVar etc. dbSNP is the most extensive among all the databases. By release of 135 hosting number of human SNPs reached more than 50 million, including 535,660 synonymous and 873,308 non-synonymous SNPs [1]. Only the non-synonymous SNPs (nsSNPs), also called as missense variants are particularly important as they result in to changes in the translated amino acid residue sequence. It is likely that nsSNPs play a major role in the functional diversity of coded proteins in human populations and have been linked with many diseases. nsSNPs may affect the protein function by reducing protein solubility or by destabilizing protein structure and they may affect gene regulation by altering transcription and translation all in ways that may not be identified by structure or phylogeny-based features [2], [3], [4], [5].
It is estimated that breast cancer may affect one out of every eight women at some point in her lifetime. Only 10% of women have a hereditary predisposition to breast cancer. Meanwhile, less than half of the patients have been found to carry a mutation in the BRCA1 or BRCA2 gene [6]. The disease may occur due to mutations in the code for the genes of the proteins that interact with BRCA1 and BRCA2. BARD1 is one of these genes and encodes the BRCA1-associated RING domain protein (BARD1). BARD1 is a protein with 777 residues. It contains an amino-terminal RING domain (residues 46–90), three ankyrin repeats (residues 427–525) and two carboxy-terminal BRCT domains (residues 616–653 and 743–777). It also has nuclear export and localization signals (residues 102–120 and the residues after 177, potentially residues 204–209) [7]. BARD1 makes a stable heterodimer in association with BRCA1 [8]. Many mutations have been identified in BARD1 in non-hereditary site-specific breast and breast/ovarian cancer cases [9], [10]. The majority of breast cancer cases (approximately 70%) are considered sporadic in nature because they do not have extensive familial history [11]. In most of these cases, BRCA1 and BRCA2 are rarely found mutated. In contrast, both germline and somatic BARD1 mutations are found in sporadic breast, ovarian and uterine cancers [12].
A somatic mutation (Val695Leu) and a germline mutation in BARD1 associated with sporadic breast cancer (Val695Leu) and one (Gln564His) associated with ovarian cancer have been reported [12]. Three SNPs namely, Lys312Asn, Cys557Ser and Asn295Ser have been found associated with BRCA1 and BRCA2 mutations in negative familial breast/ovarian cancer [9]. In spite of those findings, the functional role of BARD1 in cancer susceptibility is unclear. However, many SNPs have been reported in BARD1 but only two have been suggested to be involved into breast cancer susceptibility. Val507Met is considered to be responsible for high risk of postmenopausal breast cancer and Cys557Ser for familiar breast cancer [10], [13]. In addition of the female specific cancers, BARD1 SNPs have been found to be associated with neuroblastoma. As a matter of fact, SNPs in BARD1 coding region cause the expression of an oncogenic isoform and that influence the neuroblastoma susceptibility and oncogenicity [14] (Bosse et al, 2012). BARD1 seems a plausible target for female-specific cancer and other cancer studies. However, knowledge about the clinical relevance for many of the BARD1 SNPs is still limited [9], [10], [12]. This study was undertaken to explore and extend the knowledge related to the effect of SNPs on the stability and function of the BARD1 gene.
Results and Discussion
The dbSNP database contains both validated and non-validated polymorphisms. In spite of this drawback, we opted to avail the dbSNP because allelic frequency of most of nsSNPs of BARD1 has been recorded there (except 12 out of 50) and that is the most extensive SNP database [15]. At dbSNP, BARD1 gene contains data for 1709 SNPs. Out of 1752 SNPs, 50 are nsSNPs and 14 are in UTRs. There are 6 SNPs in the 5′ UTR and 8 SNPs in the 3′ UTR. Our investigation accounted for the nsSNPs in the coding region and the 5′ and 3′ UTR SNPs. A graphical representation of the distribution of SNPs in the coding region and the UTRs is depicted in terms of percentage in Figure 1; 2.85% of the total numbers of SNPs are nsSNPs present in the coding region, whereas only 0.34% and 0.45% of the total number of SNPs are in the 5′ and 3′ UTRs, respectively.
Deleterious nsSNPs predicted by SIFT
The sequence homology-based tool SIFT was used to determine the level of conservation of a particular amino acid position in a protein. SIFT has been tested on many human SNP databases and was found able to distinguish the disease associated SNPs from a neutral one with only a 20% false positive error. The sensitivity of SIFT is confirmed by the subset of nsSNP from dbSNP predicted to affect function were involved in disease. Furthermore, The SIFT algorithm works mainly sequence for prediction while that performs similarly to tools that use structure. Since, SIFT can predict a large number of a substitutions, as that do not requires the structures. Seventy four percent (74%) of nsSNPs identified by the SNP Consortium, were sufficiently similar to homologs in protein sequence databases for SIFT prediction. Hence, using SIFT is advantageous over other tools [16].
A .txt file containing "db SNP rsIDs" of all 50 nsSNPs was submitted to the "SIFT dbSNP rsIDs" page (http://sift.jcvi.org/www/SIFT_dbSNP.html) to calculate the tolerance index. The functional impact of the amino acid substitution is inversely proportional to the tolerance index (TI). Figure 2 and Table 1 summarize the results. Out of 50 nsSNPs 11 were predicted as 'Not Tolerable' (Table 1) and had a Tolerance Index (TI) ≤0.05. The corresponding amino acid substitutions of rs143914387, rs1048108, rs71579841, rs61754118 and rs139785364 had a TI score of 0.00. The TI score was 0.01 for rs187590361 and rs13389423, 0.02 for rs111367604, 0.03 for rs146629794 and 0.04 for rs3738885 and rs4986841. The nucleotide change C→T accounted for the maximum number (four) of deleterious SNPs, followed by A→G (two). The rest of the nucleotide changes occurred only once.
Damaging nsSNPs predicted by PolyPhen
The nsSNPs involved in structural modification were determined by the PolyPhen (Polymorphism and Phenotype) program. PolyPhen software version 2.0.9 predicts the fate of the structure and function of a protein due to an amino acid change through specific empirical rules on the sequence. Input options for the tool are protein sequence, accession number or database ID/accession number combined with sequence position with amino acid variants. For sequence-based characterization of the substitution site PolyPhen uses the TMHMM algorithm, Coils2 program and SignalP program to predict transmembrane, coiled coil and signal peptide regions of the protein sequences. PolyPhen identifies homologues of the input sequences via a BLAST and calculates position-specific independent count (PSIC) scores for every variant and estimates the difference between the variant scores, the difference of >0.339 is detrimental. The program carries out a BLAST query of a sequence against a protein structure database (PDB and PQS) for mapping of the substitution site to known protein 3-dimensional structures. PolyPhen uses the DSSP database to obtain secondary structure and solvent accessible surface area for the mapped amino acid residues. There are certain empirical rules applied on the sequences and the accuracy of that is approximately 82% with a chance of 8% false positive prediction [17].
The protein accession number of BARD1 (Q99728) and the amino acid substitutions corresponding to each of fifty nsSNPs were submitted separately. Table 2 summarizes the results obtained from the PolyPhen server. A position-specific independent count (PSIC) score difference was assigned using the categories 'probably damaging' (2.00 or more), 'possibly damaging' (1.40–1.90), 'potentially damaging' (1.20–1.50), 'borderline' (1.00–1.20) and 'benign' (0.00–0.90). Twenty-seven out of 50 nsSNPs were predicted as 'damaging,' and the PSIC scores fell into the range of 1.51 to 3.41. Five nsSNPs predicted to be deleterious by the SIFT program were also predicted to be 'damaging' by the PolyPhen server. rs139785364 had a SIFT TI of 0.00 and a PolyPhen PSIC of 2.495. Therefore, the relevant mutation would be important when manifesting itself in the cancers caused by the nonfunctioning of the BRCA1-BARD1 complex.
Functional SNPs in untranslated regions (UTR) predicted by FastSNP
The polymorphisms in the 3′ UTR affect gene expression during translation of mRNA while the polymorphisms in the 5′ UTR influence RNA half-life by altering polyadenylation [18], [19]. The FastSNP (Function Analysis and Selection Tool for Single Nucleotide Polymorphisms) program was used to predict the functionally important SNPs in the 3′ and 5′ UTRs. That is a web server that efficiently identifies the functional SNPs. That prioritizes SNPs according to twelve parameters (phenotypic risks and functional effects), such as changes to the transcriptional level, pre-mRNA splicing, protein structure, etc. FastSNP is unique in its feature that the prediction of functional effects is always based on the most up-to-date information. FastSNP extracts updated information from eleven external Web servers. FastSNP also provides project management services for registered users to store and export their candidate SNPs and update the SNPs putative functional effects by re-submitting the query [20].
The FastSNP search was performed by querying by gene symbol (BARD1). Table 3 lists the SNPs in the UTRs and the intronic region. The SNP rs58253676 in the 3′ UTR may have splicing regulator and enhancer functions and may possibly be a splice site. Most importantly, the nucleotide change may have a medium-high (3–4) level of risk for being a splicing regulator and a low-medium (2–3) level of risk for enhancer functions. rs17489363 and rs17426219 in the 5′ UTR may alter the transcriptional binding site. In the intronic region, rs67822872 SNP, an intronic enhancer, may have a medium-high (3–4) level of risk upon nucleotide change.
Modeling of amino acid substitution effects due to nsSNPs on protein structure, Energy minimization and RMSD
(A) The closest related protein structures.
By using the EMBL-EBI Web-based tool PDBsum, the BARD1 gene product-related protein structures were searched. Three related protein structures, namely 1JM7, 3C5R and 2NTE, were found to share 100% amino acid sequence similarity (Table 4). 1JM7 is a BRCA1-BARD1 complex. Chain B belongs to BARD1 and has 97 amino acid residues. Chain B accounts for residues 26 to 122. 3C5R and 2NTE are homodimers. They are the stretches of BARD1 that account for residues 425 to 545 and 568 to 777, respectively.
(B) Models of substituted amino acids and their minimized energy and RMSD.
The single amino acid polymorphism database (SAAP) server http://www.bioinf.org.uk/saap/db/ is offline due to essential maintenance. Thus, we were unable to map the deleterious nsSNPs into protein structure through SAAP. 1JM7, 3C5R and 2NTE were scanned manually to identify amino acid polymorphisms. IJM7 accounted for three nsSNPs: rs71579841 (Ala40Val), rs140254589 (Asp102Asn) and rs144856889 (His116Tyr). 3C5R also had three nsSNPs: rs137988817 (Asp458His), rs111350417 (Val477Ala) and rs149839922 (Leu480Ser). However, 2NTE had 10 nsSNPs. nsSNPs found in 1JM7, 3C5R and 2NTE are listed in Table 5. All the functional nsSNPs predicted using the SIFT and PolyPhan tools and present in the three structures mentioned above were subjected to the SPDBV mutation tool. A model for each functional nsSNP was made and visualized as a comparison using SPDBV.
Energy minimization for all the models and their native structures was achieved using the NOMAD-REF Gromacs server. The Gromacs tool uses a force field for energy minimization. The total energy for all the mutant and native models after minimization is listed in Table 5. The total energies for the native structures of 1JM7, 3C5R and 2NTE are −5209.592 kJ/mol, −6174.53 kJ/mol and −12127.86 kJ/mol, respectively. Change in total energy due to mutation is noticeable in the 1JM7 mutant rs71579841 (Ala40Val), being −2218.149 kJ/mol. Change in the total energy due to mutation is also noticeable in the 2NTE mutants rs76744638 (Arg731Gly) and rs139785364 (Arg751Try), being −11862.29 kJ/mol and −11881.313 kJ/mol, respectively. Interestingly, other mutant models had almost the same energy as their native structures.
RMSD is the measure of the deviation of the mutant structures from their native configurations. Higher the RMSD value, the more deviation between the two structures. Structure changes, in turn, affect functional activity. Among all the 16 mutants, rs144856889 (His116Tyr) had the highest RMSD (1.8039 Å), followed by rs137988817 (Asp458His) (1.1598 Å). rs140254589 (Asp102Asn), rs140729292 (Ala721Thr), rs140642433 (Cys628Arg), rs149839922 (Leu480Ser) and rs150121935 (Asp710Val) had 0.6960 Å, 0.6725 Å, 0.5496 Å and 0.5399 Å RMSD scores, respectively. Scores for other mutants fall in the range between 0.1364 (rs71579841, Ala695 Val) and 0.4724 (rs76744638, Arg731Gly). RMSDs for all the mutant structures are listed in Table 5.
Prediction of change in stability due to mutation
The I-Mutant 2.0 server was used to predict the change in protein structure stability due to mutations. the input option for this tool is the 3D structure of protein. The tool was developed and tested with the data extracted from ProTherm which is the most comprehensive available database of thermodynamic experimental data of free energy changes of protein stability due to mutation. Hence, that efficiently predicts whether a protein mutation affects the stability of the protein structure or not. The predictions are 80% or 70% accurate depending upon the usage of structural or sequence information, respectively. The tool provides the scores of free energy change predictions calculated with the energy-based FOLD-X tool. By incorporating the FOLD-X approximation with those of I-Mutant, an precision of 93% on one third of the database can be accomplished, thus making I-Mutant a helpful tool for protein design and mutation [21].
Although the stabilities of the two nsSNPs rs144856889 (His116Tyr) and rs137988817 (Asp458His) increased, their reliability index (RI) was zero (0) and one (1), respectively. Other mutants exhibited decreased stability with an RI ranging between 9 and 3. These results are summarized in Table 6.
Mutant amino acids affect the domain structures of BARD1
The affected domains and the allelic frequency of corresponding nsSNPs are listed in Table 7. Only ankyrin ring and BRCT domains harbor the predicted deleterious mutations. All the mutations of 2NTE, except rs187590361 (Asn663Ser), were located in the BRCT domains of BARD1, whereas all the 35CR mutations were located in the ankyrin rings. Structural changes in BARD1 due to 2NTE mutations can be better understood in Figure 3.
Figure shows the differences of structure and electron cloud density between native and mutant models BARD1 Protein (PDB ID: 2NTE). Models were generated by using SPDBV (v4.0).
One of the major purposes of genetics studies is to distinguish functionally neutral mutations from those that contribute to disease. About half of the known gene lesions accounting for human inherited disease involve amino acid substitutions. Hence, to identify the nsSNPs those affect protein functions and, in turn, manifest themselves as diseases are an important issue [22], [23]. The functional effect of many nsSNPs may be neutral because natural selection will have removed mutations in essential positions. Using phylogenetic information with certain structural approaches is the basis of the assessment of these nsSNPs. Still, there is increasing evidence that the onset of many human diseases is due to mutations in the intronic regions of genes. Such mutations cause alterations in regulatory regions and the splicing process [24], [25].
SNPs are widespread throughout the genome. This fact makes them a preferred choice as genetic markers in the research on diseases and their corresponding drugs [26]. More than 1 million SNPs have been reported so far. Many of them provide a large amount of information about relationships between individuals, populations and diseases. However, the large numbers of SNPs cause a challenge for biologists and bioinformaticians [26]. Studying associations between disease risk and these genetic variations using a molecular epidemiological approach has gained much attention from scientists. The number of reported and recorded SNPs is increasing. This huge number of SNPs makes it difficult for researchers to plan costly population-based genotyping. Due to a plethora of SNPs, it is difficult to choose the target SNPs that will most likely affect phenotypic functions and ultimately contribute to disease development [24], [26], [27].
Approximately 5–10% of breast and ovarian cancer predispositions are hereditary [28] BRCA1 and BRCA2 being the most studied susceptibility genes. Mutations in BRCA1 are found in 40–50% of families with a high breast cancer risk. Among these mutation occurrences, 75–80% account for both breast and ovarian cancers [29]. Even so, a significant proportion of predisposition to breast cancer that is due to these genetic aberrations is still unanswered. This leads us to hypothesize that there must be involvement of some other susceptibility genes. Therefore, we targeted the genes encoding proteins associated to BRCA1 for study.
In this study, we have examined the BARD1 gene to analyze and identify the deleterious and functional nsSNPs using in silico methods. BARD1 is one of the BRCA1-associated proteins and the two share closely related domain structures [30]. Both have an N-terminal zinc finger domain and a C-terminal BRCT domain which had been found in many proteins. In these proteins, the domains are involved in DNA repair and cell cycle regulation. Particularly, the zinc finger domain is functionally important in the formation of the BRC1/BARD1 complex [31]. BARD1 contains three ankyrin repeats, which have been reported to be involved in transcription regulation when they are also present in other proteins [32]. Furthermore, the complex of the BARD1/BRCA1 heterodimer and CstF-50 (cleavage stimulation factor subunit 1) represses the polyadenylation machinery, presumably to prevent inappropriate mRNA processing at sites of DNA repair [33]. BARD1 also regulates the nuclear translocation of BRCA1 by preventing its export [34]. The involvement of BARD1 in TP53-independent apoptotic signaling has been reported previously. It can also function independent of BRCA1. BARD1 interacts with ankyrin repeats of BCL3 and thus is likely to modulate the activities of the transcription factor NFKB [35], [36].
Hence, nsSNP variation which causes a change in amino acid composition may result in the alteration of structural domains. For example, if there is an alteration in the ring finger domain, it may hinder BRCA1/BARD1 complex formation, reduce the stability of BRCA1, and change the polyadenylation process of mRNAs. Nevertheless, the alteration of ankyrin repeats or the BRCA1 C-terminal (BRCT) domain may lead to abnormal transcriptional or cell cycle regulation, respectively.
SIFT predicted 11 nsSNPs as deleterious, and PolyPhen predicted 27 nsSNPs as deleterious. Among them, only 5 nsSNPs were common (Table 1 and 2). They are rs4986841 (Ile653Phe), rs187590361 (Asn663Ser), rs111367604 (Val695Leu), rs13389423 (Ser728Phe) and rs139785364 (Arg751Trp). rs111367604 (Val695Leu) has been found to be associated with predisposition to breast, ovarian and uterine cancers [12], which is in agreement with our findings. SNPs in UTRs may alter transcription binding sites, splicing sites and polyadenylation of mRNAs [18], [19]. The SNPs rs58253676 in the 3′ and rs17489363 and rs17426219 in the 5′ UTRs are predicted to be involved in splice site regulation (Table 3). None of them have been studied so far in terms of their functional effects in any population. There were only three structures found in the Protein Data Bank, 1JM7, 3C5R and 2NTE, which shared 100% similarity with the BARD1 amino acid sequence (Table 4). Energy minimization, RMSD calculation and modeling of mutants were performed on the above-mentioned structures. The free energies of the mutant models of 1JM7 rs71579841 (Ala40Val), C35R rs137988817 (Asp458His), C35R rs149839922 (Leu480Ser) and 2NTE rs140642433 (Cys628Arg) and rs76744638 (Arg731Gly) decreased markedly. The minimum RMSD was calculated to be 0.1364 for 1JM7 rs71579841 (Ala40Val), while the maximum RMSD was calculated to be 1.8039 for 1JM7 rs144856889 (His116Tyr) and 1.1598 for C35R rs137988817 (Asp458His). RMSDs in the range of 0.7866 for rs140254589 (Asp102Asn) and 0.1796 for rs139785364 (Arg751Trp) (Table 5) were observed in these mutants. All five nsSNPs which were predicted to be deleterious by both SIFT and PolyPhen were found to be involved in decreasing protein stability. Four of them, rs4986841 (Ile653Phe), rs111367604 (Val695Leu), rs13389423 (Ser728Phe) and rs139785364 (Arg721Trp), were found in the BRCT domain of BARD1. This finding suggests that these four nsSNPs may decrease protein stability, hinder transcriptional regulation, and interfere with cell cycle regulation [31].
BARD1 SNPs G1743C (Cys557Ser), T2006C (Cys645Arg) and G2355A (Ser761Asn) have been identified to be associated with ovarian cancer, breast and ovarian cancer and breast cancer, respectively [9], [12]. A Finnish population study reported three synonymous and four nsSNPs. The nsSNPs were C1207G (Ser378Arg), G1592A (Val507Mat), C2045T (Arg658Cys) and G1743C (Cys557Ser). Only G1743C (Cys557Ser) was found associated with breast cancer predisposition in that study [10]. The same SNP was found associated with risk of single and multiple primary breast cancer [37]. Pro24Ser and C1207G (Ser378Arg) may jointly contribute to the susceptibility of breast cancer. Their heterozygote and homozygote are associated with decreased risk of breast cancer [38]. Recently, G1743C (Cys557Ser) has been reported for no association with the predisposition of familial breast cancer in an Australian population based case control study [39]. In a cohort based study of a French population, nine common SNPs of BARD1 including G1743C (Cys557Ser) were not shown any role as modifier of risk in BRCA1/2 mutant carriers [40]. Furthermore, BARD1 SNPs rs6435862 and rs3768716 and some known common SNPs has been found significantly associated with the aggressive neuroblastoma [41].During the last decade, approximately 12 nsSNPs have been studied in different populations for their association with the predisposition to various female cancers. Some of them are recorded in the dbSNP database for the BARD1 gene (http://www.ncbi.nlm.nih.gov/gene/580). They are rs28997576 (Cys557Ser), rs146946984 (Arg565His), rs34744268 (Cys645Arg), rs111367604 (Val695Leu) and rs142155101 (Ser761Asn) [9], [12]. rs111367604 (Val695Leu) has been predicted to be deleterious by SIFT and PolyPhen, while I-mutant also predicts its decreased stability. rs146946984 (Arg565His) has been predicted to be deleterious by PolyPhen only. Contrarily, among nsSNPs predicted damaging by SIFT and/or PolyPhen rs1048108 (Ser24Pro), rs16852741 (Gly186Ser) rs2229571 (Ser378Arg) had been reported in population based studies. Studies revealed that above three nsSNPs show no significant association with disease [42], [43]. Although, rs28997576 (Cys557Ser), rs34744268 (Cys645Arg) and rs142155101 (Ser761Asn) are well studied and published nsSNPs, however, none of the tools used for the predictions were able to predict their damaging effects. Hence, there is a need of testing the predicted nsSNPs for their functional roles meanwhile; there is also a need of improveing the web-based tools for more précised predictions. Many nsSNPs have been studied in populations but not indexed in the dbSNP database, such as Asn295Ser, Lys312Asn, Asn470Ser, Gln564His, Thr598Ile and Ile692Thr [9], [12], [13], [44]. Thus, there is also a need to update the dbSNP database accordingly.
This study concludes that with the available bioinformatics tools and the data present in the dbSNP database, four snSNPs are deleterious and likely reduce protein stability. These snSNPs are rs4986841 (Ile653Phe), rs111367604 (Val695Leu), rs13389423 (Ser728FPhe) and rs139785364 (Arg751Trp). Their presence in the BRAC domain increases the possibility of altered transcriptional and cell cycle regulation. Therefore, the probability of their involvement in disease predisposition increases. This prediction can be further tested through larger population-based studies.
Materials and Methods
BARD1 gene SNPs and their protein sequences in the FASTA format were retrieved from the dbSNP database [15], [45] (http://www.ncbi.nlm.nih.gov/SNP/), and ".pdb" files for BARD1 subunits were retrieved from the RCSB Protein Data Base [46] (http://www.rcsb.org/pdb/home/home.do) for computational analysis in this study.
Sequence homology-based prediction of deleterious nsSNPs by using SIFT
The Sorting Intolerant from Tolerant (SIFT) server available at (http://sift.jcvi.org) was used to predict the deleterious coding non-synonymous SNPs. The SIFT program can sort out the functionally neutral and deleterious amino acid changes due to SNPs in the coding regions of genes [16]. For the prediction of functional consequences on proteins due to nsSNPs, the SIFT program utilizes amino acid sequence homology and the physical properties of the proteins in combination with naturally occurring nsSNPs by aligning paralogous and orthologous protein sequences. The algorithms for the SIFT program use the latest SWISS-PROT, nr and TrEMBL databases to find homologous sequences by considering the median conservation sequence score to be 3.00. The threshold for the intolerance index is ≥0.05. Seq-Rep is the fraction of sequences that contain amino acids shown in color code: black (non-polar); green (uncharged polar); red (basic); blue (acidic). A low fraction indicates the position is either severely gapped or non-alignable and has little information.
Structural homology-based prediction of functional consequences of coding nsSNPs by using PolyPhen
The ability of the protein to interact with other molecules or to have different functions depends upon its tertiary structure [47], [48]. Therefore, analysis of damaged coding nsSNPs at the structural level is necessary to understand the activity of the protein. The Polyphen server (http://genetics.bwh.harvard.edu/pph/) was used to study the functional consequences of nsSNPs [17], [49]. The PolyPhen server requires the protein sequence or a SWALL database ID or accession number as well as the sequence position of amino acid variants. PolyPhen classifies the SNPs as "benign," "possibly damaging" or "probably damaging" based on site-specific sequence conservation among mammals, as well as their location in the three-dimensional structure of the protein molecule. The term "damaging" used by PolyPhen reflects the mutations affecting protein structure and not the loss or gain of function [50]. The protein identifier from the UniProt database for the BARD1 protein "Q99728" was submitted with the position of variation along with the wild type and mutant amino acids. PolyPhen then calculated PSIC scores for each of the two variants based on three parameters, namely, (i) sequence-based characterization of the substitution site, (ii) profile analysis of homologous sequences and (iii) mapping of a substitution site to a known three-dimensional protein structure. The PSIC score difference between the two variants elucidates the amount of functional consequences that the nsSNP exerts. The PSIC score difference is regarded to be directly proportional to the impact of a particular amino acid substitution [51].
Scanning of functional SNPs in untranslated regions (UTRs) of the BARD1 gene using FastSNP
SNPs in the UTR sites are involved in the regulation of gene expression in many ways, such as RNA transcript splicing site or transcription factor binding site alteration [52], [53]. Hence, the UTRs were also analyzed for their functional SNPs. FastSNP (http://fastsnp.ibms.sinica.edu.tw) prioritizes SNPs according to twelve phenotypic risks and putative functional effects, such as changes to the transcriptional levels and pre-mRNA splicing and protein structure. The input order for the candidate SNPs was (i) input the candidate gene using the gene symbol, (ii) input a single SNP "rsID" or a list of SNP rsIDs for batch analysis and (iii) paste the novel SNP sequence. Input of the candidate gene symbol (BARD1) was used for analysis. Finally, the 3′and 5′ UTRs were analyzed. The SNP prioritization result was a list of SNPs with its risk ranking and possible function types. Risk level is ranked as 0, 1, 2, 3, 4 or 5, which signify the levels of "no risk", "very low risk," "low risk," "medium risk," "high risk," and "very high risk," respectively.
Modeling of protein structure amino acid substitutions caused by nsSNPs, energy minimization and calculating the RMSD
(A) Finding the closest related protein.
The EMBL-EBI Web-based tool PDBsum (http://www.ebi.ac.uk/pdbsum/) was used to find the proteins related to the BARD1 gene. PDBsum provides an at-a-glance overview of every macromolecular structure deposited in the Protein Data Bank (PDB). It performs a FASTA search against all sequences in the protein data bank (PDB) to obtain a list of the closest matches. The FASTA sequence of the BARD1 protein was provided in the query space. We selected only the three closest matches, namely the solution structure of the BRCA1/BARD1 ring-domain heterodimer (PDB ID 1JM7) [8], the crystal structure of the BARD1 ankyrin repeat domain (PDB ID 3C5R) [54] and the crystal structures of the BARD1 BRCT domains (PDB ID 2NTE) [55].
(B) Modeling amino acid substitution, energy minimization and RMSD calculation.
Swiss-PDBViewer (v4.04) was used to generate the mutated models of each of the selected PDB entries for the corresponding amino acid substitutions. Swiss-PDBViewer allows browsing through a rotamer library to change amino acids. A "mutation tool" was used to replace the native amino acid with a new one. The mutation tool facilitates the replacement of the native amino acid by the "best" rotamer of the new amino acid. The ".pdb" files were saved for all the models. The NOMAD-Ref Gromacs server was used to perform energy minimization for all the native and mutated models of 1JM7, 3C5R and 2NTE. The NOMAD-Ref Server makes use of Gromacs using force fields for energy minimization according to the steepest descent, conjugate gradient or L-BFGS methods [56]. The conjugate gradient method was utilized in this study. RMSDs between the native structure and each mutant were calculated using YASARA [57].
Predicting the change in stability due to mutation
To predict the change in the stability of the protein upon mutation, a support vector machine (SVM)-based tool server, I-Mutant 2.0, was used. This tool automatically predicts protein stability changes upon single point mutations. Prediction can be performed using either protein structure or sequence. I-Mutant 2.0 can be used both as a classifier for predicting the sign of the protein stability change upon mutation and as a regression estimator for predicting the related change in Gibbs-free energy (ΔΔG) [21]. Scanning of nsSNPs for their position in different protein domains
To find the nsSNPs and the amino acid changes they may cause in different domains of the protein structures, the Prosit-ExPaSy tool was used (http://prosite.expasy.org/). The UniProtKB ID was provided for the query column, and the UniProt database was searched for motifs and domains of BARD1. The results were obtained as the categorized sequence of amino acids with their respective positions in the protein subsequences and domains.
Author Contributions
Conceived and designed the experiments: AAA TNH BLG. Performed the experiments: TNH NAS. Analyzed the data: AAA TNH GS. Wrote the paper: TNH AAA BLG.
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- 36. Dechend R, Hirano F, Lehmann K, Heissmeyer V, Ansieau S, et al. (1999) The Bcl-3 oncoprotein acts as a bridging factor between NF-kappaB/Rel and nuclear co-regulators. Oncogene 18: 3316–3323.
- 37. Stacey SN, Sulem P, Johannsson OT, Helgason A, Gudmundsson J, et al. (2006) The BARD1 Cys557Ser variant and breast cancer risk in Iceland. PLoS Med 3: e217.
- 38. Hou WK, Xu YX, Yu T, Zhang L, Zhang WW, et al. (2007) Adipocytokines and breast cancer risk. Chin Med J (Engl) 120: 1592–1596.
- 39. Johnatty SE, Beesley J, Chen X, Hopper JL, Southey MC, et al. (2009) The BARD1 Cys557Ser polymorphism and breast cancer risk: an Australian case-control and family analysis. Breast Cancer Res Treat 115: 145–150.
- 40. Spurdle AB, Marquart L, McGuffog L, Healey S, Sinilnikova O, et al. (2011) Common genetic variation at BARD1 is not associated with breast cancer risk in BRCA1 or BRCA2 mutation carriers. Cancer Epidemiol Biomarkers Prev 20: 1032–1038.
- 41. Capasso M, Devoto M, Hou C, Asgharzadeh S, Glessner JT, et al. (2009) Common variations in BARD1 influence susceptibility to high-risk neuroblastoma. Nat Genet 41: 718–723.
- 42. Zhou X, Han S, Wang S, Chen X, Dong J, et al. (2009) Polymorphisms in HPV E6/E7 protein interacted genes and risk of cervical cancer in Chinese women: a case-control analysis. Gynecol Oncol 114: 327–31.
- 43. Ryu GM, Song P, Kim KW, Oh KS, Park KJ, et al. (2009) Genome-wide analysis to predict protein sequence variations that change phosphorylation sites or their corresponding kinases. Nucleic Acids Res 37: 1297–1307.
- 44. Gorringe KL, Choong DY, Visvader JE, Lindeman GJ, Campbell IG (2008) BARD1 variants are not associated with breast cancer risk in Australian familial breast cancer. Breast Cancer Res Treat 111: 505–509.
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- 46. Dutta S, M Berman H, F Bluhm W (2007) Using the tools and resources of the RCSB protein data bank. Curr Protoc Bioinformatics Dec;Chapter 1: Unit1.9.
- 47. Hasan TN, Grace BL, Masoodi TA, Shafi G, Alshatwi AA, et al. (2011) Affinity of estrogens for human progesterone receptor A and B monomers and risk of breast cancer: a comparative molecular modeling study. Adv Appl Bioinform Chem 4: 29–36.
- 48. Alshatwi AA, Hasan TN, Syed NA, Shafi G (2011) Predicting the possibility of two newly isolated phenetheren ring containing compounds from Aristolochia manshuriensis as CDK2 inhibitors. Bioinformat 7: 334–338.
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Michelle Williams: Quadricycle with Matilda Ledger!
Michelle Williams and her adorable 4-year-old daughter, Matilda Ledger, take a fun quadricycle ride around Toronto, Canada with a few friends on Saturday (August 14).
The 29-year-old actress has been in town shooting the indie dramedy Take This Waltz with Seth Rogen and Sarah Silverman. Pic centers on a young woman (Williams) whose struggles with infidelity leads her to the realization that she may be addicted to the honeymoon period of her relationships.
Earlier in the day, the mother-daughter duo was seen pulling a cute red wagon through town. 10+ pics inside…


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Author: Blackwater Fever
Publisher: not published 1915.
Description: This is a bound collection of offprints containing 16 papers in different formats. Authors include Caradanatis, Mannaberg, F W W Stephens (5x), Whipple, Deeks, Woldert (3x) Yorke (2x) Ziemann, etc, George Nuttall bookplate and Molteno Institute bookplate to endpaper. Handwritten table of contents has heading "Papers on (Malaria) Blackwater, 1901 - 1915, Authors C-Z", the collection contains many tables and folding charts. One Woldert essay is entitled: "The Opinion of Doctor Robert Koch as to the Cause of Black Water Fever" with Woldert inscription to title. The most substantial paper is the "Isthmian Canal Commission report on Hemoglobunic Fever, A Study of its Etiology and Treatment" (1911) which runs over 170 pages and is included
Order No: PIP 121099
Language: English
Price: £550.00
This book has been catalogued with the following subject terms: Tropical Medicine
Click here to see a list of titles which share these subjects and here to see a list of titles which come from the same acquisition.


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Because most cigar smokers are pretty set on their favorite brands, it's almost better to ask which coffees go with your cigars. In reality, any coffee goes well with cigars and vice versa. Tobacco and coffee have tastes that naturally complement one another. If you want to find the perfect match for you, consider the following ideas. Sometimes, one of the best cigar accessories you can have is a French press or espresso maker.
First, try to match your cigar to your coffee in terms of shade. The mellow flavor of some of the darker wrapper cigars goes well with espresso drinks, provided that they don't have a lot of milk in them. If you do have a lot of milk in a drink, consider smoking a cigar with a lighter wrapper with that coffee. It will add a sharper taste to the coffee and cigar experience and can make both downright refreshing.
The standard cup of Joe goes with just about any cigar. You can pair them based on when you're having the combo, however. Some people like a strong cigar with their morning coffee. The elevated nicotine levels pack a jolt and, combined with the coffee, you'll be ready and raring to go.
If you're unwinding after dinner, consider a darker coffee to go with you cigars. You'll still get the jolt from the caffeine, but it's more subdued. Remember that lighter beans sometimes have more caffeine in them because they aren't roasted for long.
This can make them excellent for the morning, but a nice French roast is excellent after dinner, particularly if you don't' like wine or brandy, the more traditional accompaniments to a fine cigar. Your cigar will balance out the taste nicely and they both leave a great aftertaste in your mouth.
You can combine virtually any cigar and any coffee. If you have a cheaper cigar, sometimes a cup of coffee will make it taste better because it brings out the better flavors of the coffee. Likewise, a weak cup of coffee can be made more palatable with a cigar accompanying it.
Now remember that smoking strong cigars and drinking strong coffee at the same time can produce quite a buzz. If you're not used to it, take it easy. You can easily end up with a serious case of the jitters if you overdo it and, because of that, you will want to make sure you take into account that both nicotine and caffeine are stimulants and you'll want to dial your intake of both a back a little when you're having both at once!The Art of Pairing Cigars and Coffee by David Sabot


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The The Bistro Edwinstowe pub is located in central Edwinstowe. We always have something special on offer: when it's cold outside, keep warm by the warm atmosphere while in summer you will enjoy our outdoor seating area.
From a pint to a cocktail, from a sandwich to a business lunch, we offer such great prices you'll keep coming back for more! We look forward to welcoming you, whether you're having a snack at lunchtime or coming in for a drink with friends in the evening! Check out our latest promotions and the menu on our new website.
The Bistro Edwinstowe
19 High Street
Phone: 01623 824 880
|12:00 - 15:30
|Tuesday - Saturday
|18:00 - 21:00
Every month we will offer new event...
Please check what new ofer each month...
We promise you will enjoin...


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Well I came across the HiyaHiya brand of inter-changeables and fell in love.
Unlike my other brand the join is seamless, they are metal, the cable swivels and the case which contains the tips and the cables is beautifully designed. It is so compact as to be completely practical to throw in your luggage and always be prepared for impromptu yarn purchases.
I'll be casting on now.....getting in some zen time before starting the master class in a few hours.


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Willmar Hand Rox Another Loss
The St. Cloud Rox lost 7-5 Sunday night at Willmar to suffer their 8th straight loss. The game was shortened to 5 innings due to rain. Willmar scored 6 runs in the 3rd inning and St. Cloud responded with 4 runs in the 4th inning. Carlos Lopez went 2-2 with 2 runs scored, Mark Rhine went 2-2 and Boo Vazquez was 1-2 with 2 RBIs for the Rox. Paul Schaak threw 3 innings with 6 hits and 7 earned runs allowed to take the loss.
St. Cloud is 5-13 and will host Willmar tonight at 7:05, pregame on AM 1390-the Fan at 6:35. Tonight is Super Hero Night at Joe Faber Field. Fans are encouraged to wear their favorite Super Hero gear.


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What is the connection between designing services and urban and regional development? What are the competencies that the service designer can make available to policy makers and citizens?
Territorial transformation involves a process that brings together various actors: policy makers, associations, public institutions, businesses, and citizens. The change goes through the adoption of new technologies, new processes, new skills to make the places where we live "smarter", connected, sustainable, more focused on citizens' needs. In this context, service design can help to innovate the public sector, considered as a complex system of services offered to citizens, as well as to contribute to design and prototype new public policies. The emerging need to renew relations between citizens and the government goes hand in hand with redesigning public services in their organizational dimension, through public spaces, in the new dialogue between civil service and citizens, and between institutions themselves. The relationship between service design and policies has been investigated by the Department of Design of Politecnico di Milano through research activities like the Design for Europe platform and the Design Policy Lab.
In the Specializing Masters in Service Design, organized by POLI.design, students are asked to address public service issues, exploring topics such as health and, on another level, imagining the contribution service design can make to creating new policies through participatory processes and co-design activities. The fifth edition of the Masters will start on October 10, 2017.
More information available at http://www.polidesign.net/en/servicedesign


text: My teenage son is now almost 4 yrs post BMT for ALD. He continues to be well and very active with stable MRI's, he copes very well with his Addison's. He has been experiencing nightmares/bad dreams about how things might have been if he had not had a BMT. They are always the same... He falls out of his wheelchair and people just walk past him, no one stops to help. Does anyone else experience similar? Could counselling help? Thank you in advance.


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|Resource Name &
Link to Resource:
Relentless Pursuit of Success
|Captures the essence of Yu Minghong's personal philosophy towards life and business. Illustrates the lessons he has learned in life and how he has applied them to his corporate success in modern China. Yu Minghong. Cengage Learning Asia Pte. Ltd. Vol. 7 of the Asia Learning Collection. Available through the Gale Virtual Reference Library (GVRL).
|Resource Type(s):
|Access information:
|Resource is available on-campus or off-campus via EZProxy; Click the resource name to access it or here for off-campus instructions.
|Fulltext is available.
|Vendor Name:
|Gale Cengage Learning
|Research Guides:
|Business, Philosophy
|2007 ed.
|Special usage notes:
Please note: To limit your search to the contents of this specific title, select within this publication in the Quick Search box or choose Limit the results by publication title in the Advanced Search. Otherwise, you will be searching some 150+ different resources in the GVRL.


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ITS provides and supports a variety of conferencing solutions to help you collaborate with colleagues, whether they are located across campus or across the world.
- Audiovisual (AV) Service
ITS provides AV service for conference rooms on the UM-Ann Arbor campus, helping units design and build cost-effective solutions that are integrated with university infrastructure.
- BlueJeans
BlueJeans provides cloud-based conferencing with audio, video, and content sharing.
- Google Hangouts
Use Google Hangouts to chat, make voice and video calls, and share your screen. Hangouts are synced across devices, so you can start a Hangout on your computer and continue on another device.
- Adobe Connect
Adobe Connect is a web conferencing system that supports distributed meetings and training sessions with audio and video. It also allows desktop sharing, presentations, polling, and many more features.
- Campus employees whose units are members of the University of Michigan IT Asset Management (ITAM) also have access to Adobe Connect. ITAM customers can contact email@example.com or log in to the ITAM SharePoint Site for assistance or questions.
- Videoconferencing Room Systems
ITS makes videoconferencing easy with a range of videoconferencing room systems and services tailored to your unique needs—consultation, installation, training, and ongoing support. We can also register and/or support your unit's existing videoconferencing system.
- Videoconferencing Robots
ITS is investigating the benefits of videoconferencing robots. If your unit is interested in this new technology, we can consult, recommend brands, and connect unit robots to the university network.
If you have accessibility needs for videoconferencing:
- Students should contact the Knox Center for more information.
- Faculty & Staff should review Change in the Ability to Work for information.


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Legendary filmmaker Miklos Jancso, Jr. returns to the screen with this loose, improvised meditation on the current state of his native Hungary. Kapa (Zoltan Mucsi) and Pepe (Peter Scherrer) are the pair of hard-drinking, fast-talking gravediggers from his previous Peter Scherrer (1999). This time around, the two...more
All Anyad! A Szunyogok cast + crew photos
Movies.com, the ultimate source for everything movies, is your destination for new movie trailers, reviews, photos, times, tickets + more! Stay in the know with the latest movie news and cast interviews at Movies.com.
Copyright © 2019 FANDANGO MOVIES.


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Southern Fried Sunday Mustard Seed Benefit/All up and down Mills
Southern Fried Sunday Mustard Seed Benefit
The Kidney Stones, Lone Wolf OMB, The Bloody Jug Band, Kentucky Knife Fight, Andy Matchett & The Minks, The Applebutter Express, The Mud Flappers, River Bottom Nightmare Band, Hindu Cowboys, Lauris Vidal
Will's Pub
1042 N. Mills Avenue
Orlando, FL, 32803
This event is 18 and over
The Wholetones
Formed in 2007 when Alex Dorris and Taylor Freydberg met at a local bluegrass jam, the Wholetones were later completed with the addition of Russ Depa on upright bass, and Mayo Coates on drums.
They play a unique mix of bluegrass, folk, jazz, and metal, all translated through acoustic instruments.
Combining impressive technical skill with the spontaneity of jazz and drive of bluegrass, they put on ...a captivating live show, jumping from folk to metal as easily as they do from banjo to cello.
The Wholetones' blend of genres is a result of the four members' enormously varied musical appetite. They play and listen to reggae, metal, hip-hop, bluegrass, and everything in between, and the end product is a truly original sound.
The Kidney Stones
The Kidney Stones started out just like any other band; sitting in the pub at 4am, talking about accordions and St Patrick's Day 2006. Coming from metal, ska, jazz and of course punk rock genres, an Irish pub band was the obvious solution.
The band lasted 2 years before realizing it's potential as a genuine rock band, writing its own songs and playing more than just Irish drinking songs. The original music that spawned from the Kidney Stones in these early years melded the framework for the bold, outrageous, Guinness-soaked music that still sprouts from the Kidney Stones today.
The bands influences are once again typical; ranging from the Pogues, Pantera, Hepcat, Fear, Gypsy Kings, Rage Against the Machine and of course the great Neil Diamond. With all these influences, anything can come out of the Kidney Stones at any time. A Viking metal intro will turn into a beer slinging dance party. French accordion waltzes turn into southern rock. Irish jigs turn into, well, louder irish jigs!
On the 5 year anniversary of the band, they decided to add another key member to the band. It just so happens that the drummers soon to be wife is an amazingly talented fiddle player. It was too perfect to pass up. The Kidney Stones are now complete with a lineup of fiddle, accordion, harmonicas and of course guitar, drums and bass.
The Kidney Stones are now in their 7th year as a band, and don't plan on stopping anytime soon. Coming soon to a pub near you.
Lone Wolf OMB
Lone Wolf is a one man band who consist of Bruno Esposito singing raspy, dirty, vocals, playing slide banjo, bass drum with his left foot and giving a thumb tap rhythm against the banjos snare like drum, while picking the strings and playing harmonica. This style creates a unique, real, authentic sound that is unlike anything you"ve ever heard.
Founded in 2009 and hailing from Orlando, Florida, this 8-piece group has adapted traditional Jug band instruments like washboard, washtub bass, mandolin and harmonica to create their unique sound. The band has performed all over the South Eastern United States and shared the stage with notable acts such as Reverend Peyton's Big Damn Band, Joe Buck, Edwin McCain, 3 Bad Jacks, Old Man Markley and many others...
One day, an interdepartmental love affair between the SIUE school of Art, the Music Department, and the School of Education led to the conception of a 4 man dirty rock group. Nate Jones, Jason Holler, James Baker and Jason Koenig had either shared beers, classes or conversations prior, and after a few songwriting sessions and a few more parties, they decided to harness their collective creativity.
Originally starting off as a folk/rockabilly hybrid, the group underwent a dramatic and loud change over a small amount of practices, and was born as a bluesy garage rock outfit on the stage of Stagger Inn during an open mic night. Armed with only 3 songs and 15 watt practice amps, the small, not quite appropriately dressed band slayed the crowd, and the name "Kentucky Knife Fight" quickly circulated amongst the college town dwellers. Months later, the band added a close friend, David Wiatrolik, to the fold as a second guitarist.
The combination of Holler's background in poetry, Baker's and Jones's intuitive self-taught musicianship, and Wiatrolik and Koenig's strict and finely crafted Jazz leanings have led to songs that show great maturity since that first Open mic night years ago. Even the songs written for their debut LP "The Wolf Crept, The Children Slept," which has gained critical praise in Midwestern circles, have been surpassed by newer and as yet unrecorded material. The twang is twangier, the dirt is dirtier, the shifts more dynamic, the bluegrass, blues and punk influences more integrated, leaving their respective genres to become something else entirely.
Andy Matchett & The Minks
Andy Matchett & The Minks are from Orlando, Florida... Yeah, let's just get that out of the way right now - ORLANDO - A tourist town built on the fine arts of pop music & cartoons. How can meaningful rock music come out of that much sunshine and suburban happiness? The truth is, the dichotomy & secret suffering of the City Beautiful is as important to Andy Matchett's songwriting as any cold, northern borough could ever be. Just like their hometown, The Minks are known for the good times - a live show that "completely transforms the sphere of interaction" with confetti blasts, leaf blowers & a gigantic parachute that covers the crowd - sure, its a tourist attraction. But, when you really get to know the city - and the band - there's a darker, more cultured undercurrent just beyond the fun that's absolutely begging to be explored. Case in point, The Minks' debut album "The Apple Tree Circle" has been hailed by both local fans & critics alike as "near perfect", "truly exceptional" & "a universal pleasure". As much as AM/TM bring the party at shows, the focus in the studio is on seasoned songcraft and lyrical depth. From tongue in cheek "party" tracks like "Too Much Happiness" & "Thoughtsalot" to the emotionally nude ruminations of "Blind Owl" & "(I'll See You On) Monday", this band is as truly interesting & sincere as it gets. As you comb through the crowded landscape of bar bands, mainstream douche rock & hipster fashion trends, Andy Matchett & The Minks stand out as a true original. A band equally at home in the glittery limelight & down in the dumps, existing somewhere between indie & mainstream, party & patience, theme park & Winter Park... all in the image of the wonderfully misunderstood city it came from.
The Applebutter Express
Some people may discover The Applebutter Express through their new song "Hey My Brotha," a single from their sophomore project One For The Home Team, which is featured in the Ron Howard film "The Good Lie" starring Reese Witherspoon released in October of 2015.
However, the band's story begins in 2004, when Kyle Biss met his future wife Shannon while working at a record store in Bradenton, Florida. Still a focused bass player, Kyle picked up a ukulele in 2010 and began to write songs, with the idea of singing with Shannon (his then girlfriend) for fun on the couch. Pretty soon the duo began making a name for themselves among the open mic community; in no time, the eclectic, tuneful duo – now known as The Applebutter Express – were the talk of Tampa, Fl. Kyle and Shannon married in 2011, and in 2012 added Joe Trivette on the fiddle. Newest member, Zach Rogers (bass) joined in 2015.
Since then, The Applebutter Express has performed at such prestigious festivals as Bonnaroo Music and Arts Festival, Hulaween, Suwannee Springfest, and many others.
Besides their newest, One For The Home Team, Applebutter Express has released one prior full length album; the prolific foursome has a third album already in production.
Brandishing a wide arsenal of traditional acoustic instruments, this seven piece mini-orchestra has developed a reputation for ecstatic live performances and infectious onstage energy. Pulling influences every which way from the depression soaked blues of the dustbowl to the prohibitionary jazz of your local speakeasy. This sonic gumbo will have you coming back for seconds every time!
Now in their 17th year of performing together The Hindu Cowboys continue to serve as a Central Florida's veteran troubadours of Americana music.
The Hindu Cowboys sound is rich with folk, traditional string band and corn-fed roots music, all topped off with retro-country gravy. Their live sets are often laced with cover songs from old-school country (pre-1976) and classic rock, reinvented in the Hindu Cowboys style. The band frequently tours and performs as a 4-6 piece band, but principally has a writing and arrangement core based around its three founding members: Joseph Martens, playing acoustic guitar and lead vocals, Craig Roy on upright bass, and George Dimitrov on electric guitar. The musical style of the Hindu Cowboys is a unique combination of the musical roots of the origins of founding members, from Nashville, to the Southwest corner of New York State, and across the ocean to Bulgaria. The current lineup adds some of the best musician talent from Central Florida, including Gary 'Doc' Hibbard on pedal steel, fiddle, mandolin and electric guitar; Jessy Lynn Martens on fiddle, mandolin and vocals; and Gabriel Collazo on drums and war paint.
The Hindu Cowboys were formed in 1998 in Orlando, Florida, and still make Central Florida their home. The band released its debut seven-song independent album in 1999, which received world-wide critical acclaim and airplay. The attention of their first CD and their live performances caught the eye of The Wildhorse Saloon (in its heyday, it was a gateway to the Nashville labels) earning them week-long showcases at their venues in both Orlando, FL and Nashville, TN.
In 2004, the band released "Live After Five, the Hindu Cowboys; second album, crafted as a tribute to the band's loyal fan base. The record contained fifteen (15) favorites from the Hindu Cowboysplaylist.
The band's music made its way into the movies in 2006, when" Alabami Bound", a cut from the band's first album, was added to the soundtrack for Altered - a sci-fi movie by the director of The Blair Witch Project.
The band released its third CD in December, 2007 - a five-song EP which is a prelude to a long-awaited return to studio recording.
The Hindu Cowboys perform at many notable gatherings annually that includes Great Blue Heron, Riverhawk, String Break and the Albino Skunk music fesitvals. They made an appearance on the WARPED Tour during the tour's 2002 Central Florida stop as well as appearing on the main stage of Orlando Calling 2011.
The Hindu Cowboys have shared the stage with Dale Watson, Peter Rowan, Donna The Buffalo, Fred Eaglesmith, Drivin' n' Cryin', Jim Lauderdale, Marty Stuart, Los Lobos, The V-Roys, Chris Isaak, The Flatlanders, Dwight Yoakam, Restless Kelly, Del McCoury, Steep Canyon Rangers, Warren Haynes Band, Hank Williams III, The Waybacks, Tim O'Brien, Rhonda Vincent, The Gourds, The Iguanas and Robert Earl Keen, among many others.
Lauris Vidal
Lauris Vidal has always treated the neck of his guitars like the skate parks of central Florida, where he grew up. Lauris has navigated both instrument and skate session with an aggression and ease of stride that serves not to impress or intimidate, but to inspire and invigorate.
$10.00 - $14.00
Off Sale
Who's Going
Upcoming Events
Will's Pub
Fri, January 18
Southern Culture on the Skids
Sat, January 19
Brown Note
Sun, January 20
Southern Fried Sunday 13 Yr. Anniversary Mini-Fest w/ Matt Woods, Thomas & Olivia Wynn, Terri Binion, Laney Jones & More!
Sun, January 20
Honey Cutt
Wed, January 23
Camp Howard


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Sick Hero
One day when I wanted to back to my home. I felt dizzy and sick. Because result from math examination from my school. But, I still did my activities when I was sick. In middle journey to my house (likely), I found a money. But not valuable to me. Because it were 1 cent coin. So, I forgot about that. I straight to walked to my home.I felt more dizzy and more sick (I think it's ill). I was slowen my walk like snail. I went across the zebra cross and through the shop and market (Like a song from Dragonforce called "Through Fire and Flame"). I met my old friend in my old friend from elementary school. I think you knew if I was school in High School ? So I chat with my friend and I can't spoke with him about my sickness that I felt. Several minutes later (almost 15 minutes), I finished my chat with my old friend and straight walked again to my home.Suddenly, when I neared to the Money Credit Machine. I found a hole in there. I felt very VERY dizzy, sick, and confuse. I think a thief was stealing a lot of money in Money Credit Machine. I wanted to calling police but I didn't knew where I could found police station in here and I didn't knew about emergency number to calling police. So, I straight to walked to my home. Suddenly, between 2 walked road that in side were avenue, I was collide with someone in the side of avenue. I saw him and he brought a big bag. And he immediately ran straight forward. But, in his bag has a small hole. When I saw it, it were a paper and a paper is flew from his bag and it were a money !!! And he were a thief.I chased him. He were very fast to catched him. Luckly, I was brought my skateboard in my backpack. But, when I wanted to use it, in the side walked road there was police to chased a thief. I can't hold peace to chase a thief. But I felt little dizzy and sick. I went to catch a thief. Many 4 roads in there. So I followed a thief road. Luckly again, he didn't drive a car. So it were very easy catch him. And finally, I could knock out a thief with my skilled skateboard. Police were coming and catched a thief. Police thanked to me about catched a thief and Police rewarded me a 20 dollars and a medal.After the incident. I not felt dizzy and sick. I could continued my journey to home with my skateboard. About money from police I saved it to my money box and I used my money again to bought a Nintendo Dual Screens.
Silahkan copy dan paste di buku tugas anda jika anda mempunyai tugas mencari Recount Text di Internet. Karena ini merupakan karangan saya sendiri.


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Hy guys!I want to buy a book, animation book! I heard about an website animationabc, about animate that, a book for beginners, non software specific, that explains the principles of animation in depth. I don't know what to do? It's a good start for a beginner like me?
What do you think?


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The initial stages of the plan include paint, a stencil, curtains, a road side dresser, a new light fixture, pillows, accessories, a headboard and a bassinet. Oh, and patience, lots of patience!
My inspiration...
I love this fabric. Nothing fancy, just a genuine attraction.
The Old Nikki would have jumped right in and bought things without thinking things through. The new Nikki makes a plan to save money and energy...not always, but in this case, good for me:) I am looking forward to sharing our progress (psst, notice how my email went from "me" to "we" when referring to progress? Thank goodness for Henry!) I'll be in touch!


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It's been a while since I've done ATC... when I think how many I did and all the groups I used to belong to, it's weird how we change... but needless to say that I still love doing those little pieces of art. A 2½ X 3½ working surface can be very challenging!!!
These have all been done with Tim Holtz's stamps. The technique I used is one I love very much Reinker Splash (2-08) of Technique Junkie with some little additions... like peal X and ink dabbing.


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Case files.
Case files.
We can officially say now, that David Lynch has lost his mind completely :)
This has nothing to do with the old Twin Peaks I loved 25 years ago, it is almost a random series of oddities.
But don''t misunderstand me, I love it, it is just completely different.


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Premarital counseling
By KATE COLEMAN
Staff Writer
Justin Harbert says he was a little apprehensive about having to go to class to get married.
But now that the Hagerstown man and his fiancee, Susanne Jardinier, have participated in a marriage preparation program, he would recommend the program to others.
Harbert, 22, a Lutheran, says his fiancee's parents, Lucille and John Jardinier, previously led the Catholic program for the local area. He says that took a little of the edge off.


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Longines Flagship Black Dial Ladies Watch L42164526Longines
Updated at 7/19/2014 07:56:53 - Details
Stainless steel case with a stainless steel bracelet. Fixed bezel. Black dial with silver-tone hands and index hour markers. Date display appears at the 3 o'clock position. Quartz movement. Scratch resistant sapphire crystal. Case diameter: 26 mm. Deployment clasp. Water resistant at 30 meters/ 100... see all