http://2011.igem.org/wiki/index.php?title=Special:Contributions&feed=atom&limit=500&target=Pkurani&year=&month=2011.igem.org - User contributions [en]2024-03-28T21:16:53ZFrom 2011.igem.orgMediaWiki 1.16.0http://2011.igem.org/Team:GeorgiaTech/TeamTeam:GeorgiaTech/Team2012-01-13T00:43:16Z<p>Pkurani: </p>
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Dr. Eric Gaucher<br />
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Dr. Eric Gaucher exploits a multidisciplinary approach towards research to generate an 'evolutionary synthetic biology'. This combination of molecular evolution and biomedicine provides a better understanding of basic molecular processes while simultaneously generating biomolecules useful for industrial and therapeutic purposes. Dr. Gaucher received his Ph.D. in 2001 and then worked for NASA until 2003 studying the Origins and Evolution of Early Life. He was then Researcher/President of a non-profit research organization until 2008 at which time he accepted an Associate Professor position at the Georgia Institute of Technology.<br />
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Dr. Brian Hammer<br />
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Dr. Brian Hammer is a molecular microbiologist interested a process of chemical communication called Quorum Sensing, which bacteria use to orchestrate group behavior. He received his MS in aquatic ecology from the University of Michigan in 1995 and his PhD in Microbiology from the University of Michigan Medical School in 2001. He was a National Science Foundation (NSF) Postdoctoral Fellow and Research Scientist at Princeton University from 2001-2008. Dr. Hammer joined Georgia Tech in 2008 where he is currently an Assistant Professor in the School of Biology. Dr. Hammer is funded by two programs at the NSF: the Division of the Molecular and Cellular Biosciences (MCB); and the Network Science and Engineering (NetSE) program. <br />
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Many of his lab members study the aquatic pathogen Vibrio cholerae, with a focus on the molecular mechanisms that permit this bacterium to convert extracellular chemical signals into changes in the expression of genes important in marine systems and the human host. In addition, other members of the Hammer lab are developing synthetic quorum sensing systems to model, simulate, and experimentally validate the fundamental limits and protocols for molecular communication. This work is in collaboration with several professors in the College of Engineering at Georgia Tech.<br />
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Dr. Harold Kim<br />
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Dr. Harold Kim is an experimental biophysicist who studies the biophysics of the genome and its influence on gene expression patterns. His research group uses single-molecule and single-cell fluorescence microscopy to understand (1)how intrinsic DNA looping dynamics is related to nucleosome formation; (2) how nucleosome positioning influences gene regulation; and (3) how spatial distribution and copy number of genes influences their expression patterns. Dr. Harold Kim received his PhD in applied physics from Stanford University in 2004, did his postdoctoral research at Harvard University from 2005 to 2009, and joined the School of Physics in 2010 as an assistant professor. He is the recipient of a Burroughs Wellcome Career Award at the Scientific Interface.<br />
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Dr. Mark Styczynski<br />
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Dr. Mark Styczynski uses experimental and computational techniques to study the connections between the different layers of regulation in cells and the cells' ultimate phenotypic outcomes. Of most interest to his lab are metabolites, the small molecule building blocks necessary for all cellular functions both as source materials and as cues that prompt regulation and other cellular responses. He studies metabolism in a variety of systems ranging from yeast to human cancer cell lines. Dr. Styczynski received his PhD from MIT in 2007, was a postdoctoral associate at the Broad Institute from 2007 to 2009, and joined the School of Chemical & Biomolecular Engineering at Georgia Tech in 2009 as an Assistant Professor.<br />
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Dr. Joshua S. Weitz<br />
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Dr. Joshua Weitz is a quantitative biologist interested in the structure and dynamics of complex biological systems. He received his PhD in Physics from MIT in 2003 and was a NSF Postdoctoral Fellow and Associate Research Scholar at Princeton University from 2003-2006. Dr. Weitz joined Georgia Tech in 2007 where he is currently an Assistant Professor of Biology with courtesy appointments in Physics and Bioengineering. Dr. Weitz is the recipient of a Burroughs Wellcome Career Award at the Scientific Interface and is funded by the James S. McDonnell Foundation, NSF, and DARPA. His research group includes ecologists, mathematicians, physicists and bioinformaticians working on four major research themes: (i) viral dynamics at the molecular, population and evolutionary scales; (ii) quantitative systems biology and bioinformatics; (iii) structure and function of vascular networks; (iv) theoretical ecology and epidemiology. The work in the Weitz group is primarily theoretical in nature, and utilizes the tools of nonlinear dynamics, stochastic processes, and large-scale data analysis to interact with experimentalists. Examples of recent and ongoing projects include studies of collective decision making in bacterial viruses, robustness and fragility of gene regulatory networks to copy number variation, unsupervised approaches to binning short environmental sequence fragments, network phenotyping and classification of root system architecture, and a Hierarchical Bayesian analysis of allometric scaling models in biology.<br />
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Ryan Randall<br />
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"Ryan manages Prof. Gaucher's laboratory and performs original research as a 2nd year graduate student within the School of Biology at GaTech. She is currently evolving monomeric red fluorescent proteins in the laboratory. Her data will be used to construct an experimental phylogeny that will in turn be used to benchmark computational methods of ancestral sequence reconstruction. She received her BS from the University of Georgia in 2008 with a degree in Cellular Biology. She is the graduate advisor of the Georgia Tech iGEM 2011 team and has been a constant source of support for the team. She has proven invaluable to GTiGEM 2011 team and has provided us with all the lab supplies and protocols."<br />
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<br>-Mitesh Agrawal<br />
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Mitesh Agrawal<br />
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""Mitesh is a 3rd year international student from India majoring in Biomedical Engineering. Besides being all around awesome, he was a member of Georgia Tech’s first iGEM team where he was a major contributor to the modeling section of the project. This year he has once again been a vital part of the team, helping with experimental design and giving us perspective on how things worked last year. His research endeavors have included study of embryonic stem cells and genetic engineering. Mitesh was a crew member (radio specialist and biologist) for the Georgia Tech Mars Desert Research Station and he is also actively involved in the Georgia Alpha chapter of Tau Beta Pi - an engineering honors society. When not in the lab, Mitesh enjoys playing a rousing game of racquetball and reading science fiction novels. Oh, and he is a die-hard Manchester United fan, so watch out Liverpool. >:)”<br />
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<br>-Hannah Keith<br />
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Calvin Goveia<br />
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"Calvin is a 4th year chemical engineer specializing in bioprocess engineering. He is very interested in using ethanol as a fuel, and looking into the process of the utilizating of recalcitrant biomass to create renewable bioenergy. He would like to improve the current process of producing cellulose based biofuels by cloning essential enzymes from cellulolytic bacteria. He hopes to engineer E. coli with these genes to simultaneously hydrolyze cellulose and ferment those products of hydrolysis. Calvin is also working on the modeling component of our project."<br />
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<br>-Paul Sebexen<br />
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Kettner Griswold Jr.<br />
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"Kettner Griswold is a straight up mad scientist. He is a 2nd year MSE Student who hails from Bethesda, MD where he was a local legend- seriously, google him. Kettner worked at the J. Craig Venter Institue for a year during high school and now spends his days reading scientific literature and white water kayaking (usually not at the same time)."<br />
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<br>-Hannah Keith<br />
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Hannah Keith<br />
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"Hannah is a second year biochemistry major and is a vital member of the GTiGEM 2011 team. Besides being an important contributor in experimental design of this year’s project, she is the one who has kept our team organized and well run. Her field of interest is studying structural and functional properties of different biochemical molecules, including a special fondness for H2O. Hannah loves running and is an avid reader of non-fictional books. When not busy with studies and iGEM, she devotes her time to various community services and organizations like the American Medical Student Association. Overall, she has been an amazing addition to the Georgia Tech iGEM team."<br />
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<br>-Mitesh Agrawal<br />
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Priya Kurani<br />
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"Priya is a 4th year Computational Media major. She is interested in how modern media channels can help illustrate complex processes in Synthetic Biology. She likes intuitive user interfaces, Afrojack, and Cuban food."<br />
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<br>-Paul Sebexen<br />
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Paul Sebexen<br />
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"Paul Sebexen is a 3rd-year Computer Science major. He is originally from New York City and has research interests ranging from computational biology to solid-state physics. In his free time, Paul enjoys studying languages, bicycling, tennis, and playing piano. Paul loves everything German."<br />
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<br>-Priya Kurani<br />
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We received the sequence of S. thermophilus DGCC7710 CRISPR1 locus, which was graciously provided by Rodolphe Barrangou and Danisco, Inc. We also signed a nondisclosure agreement with said party. Promptly after the sequence was received we designed primers to amplify the entire locus. <br><br><br />
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Cultured cells overnight and performed a genomic DNA extraction. We used primers to amplify out the CRISPR locus via PCR. <br><br><br />
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Using the native plasmid pNT1 as a shuttling vector for the CRISPR was extremely ineffective, and we consistently got low yields of plasmid DNA. <br><br><br />
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During the adviser meeting, one of the important ideas in our CRISPR project is suggested by Dr. Hammer. It is found that bacillus subtilis which is a gram positive bacteria do not have the CRISPR mechanism present in them. Although, it is suggested that similar promoters might be present in B. subtilis as present in the Streptococcus thermophilus. So, we designed the experiments this week for introducing the CRISPR system in the bacillus subtilis1st round of Bacillus cultures arrive, are grown on LB media and agar.<br><br><br />
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Welcome to Georgia Tech iGEM 2011 Wiki!<br />
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Purpose: As a research team, our goals are two fold: to mobilize the crispr/cas bacterial immune system on a plasmid, and subsequently utilize this system as an intelligent gene targeting system capable of eliminating antibiotic resistance.<br />
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<center><b>Title:</b> De Novo Adaptation of Streptococcus thermophilus CRISPR1 Defense in E. coli</center><br />
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The ultimate goal of our project was to utilize this system as a mobile gene targeting system capable of eliminating antibiotic resistance. The protoype construct, pSTINGER, was designed for quantitative modeling, proof of concept, and vaccination on a single plasmid. As a result of experimental error, intellectual property agreements, material availability, our biobrick constructs remain experimentally theoretical, though we have rigorously modeled our system in various scenarios. <br />
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<center><b>Title:</b> De Novo Adaptation of Streptococcus thermophilus CRISPR1 Defense in E. coli</center><br />
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The ultimate goal of our project was to utilize this system as a mobile gene targeting system capable of eliminating antibiotic resistance. The protoype construct, pSTINGER, was designed for quantitative modeling, proof of concept, and vaccination on a single plasmid. As a result of experimental error, intellectual property agreements, material availability, our biobrick constructs remain experimentally theoretical, though we have modeled our system in various scenarios. <br />
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<center><b>Project Overview</b><br><br></center><br />
Clusted regularly interspaced palindromic repeats (CRISPRs) and their associated cas genes confer immunity to bacteria and archaea by incorporating segments of foreign DNA, known as spacers, into their chromosome to recognize and inactivate the invader in the future. Our research project aims to clone the CRISPR1 system from Streptococcus thermophilus strain LMD-9 and transform it into Escherichia coli and Bacillus subtilis. The design of our experiment was motivated by the increasing threat of antibiotic resistant bacterial populations to human health. By designing bacteria that have a fitness advantage from the CRISPR system, we could wipe out these dangerous bacteria under specific circumstances. The cas9 gene from S. thermophilus LMD-9 works as an endonuclease, in a manner similar to RNA interference, to recognize and cleave the antibiotic resistant plasmid from populations of bacteria using predesigned spacers, such as ones for kanamycin resistance.<br />
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<center><br><br><b>Background</b><br><br></center><br />
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It has long been known that bacteria protect themselves against threats from bacteriophage attack through a variety of methods, apparent in the existence of restriction enzymes.<br><br><br />
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Streptococcus thermophilus is a lactic acid bacteria that is widely used as a starter culture for yogurt products in the United States. All sequenced strains of Streptococcus thermophilus are known to have at least one CRISPR locus, which work with varying degrees of success. <br />
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Submitted our track selection, project abstract (which completely changes later), our team roster, and our safety questions. For our track selection, we chose 1) New Application, 2) Health/Medicine, and 3) Foundational Advance.<br><br><br />
We had a meeting with our advisers where considered our final experimental design and our final options for the biobrick. With the September 28th deadline, everyone was concerned about making the deadline to submit. Our options were to PCR cas 9, PCR CRISPR, or synthesize the repeat region. It was also discussed that we could not submit any DNA from S. thermophilus DGCC7710 due to a nondisclosure agreement for material transfer.<br><br><br />
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We placed an order for genomic DNA from LMD9 to amplify CRISPR out of it and hopefully clone it in a vector which could then be inserted into E. coli cells.<br><br><br />
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We designed primers to amplify out the entire 9kb locus of CRISPR out of the LMD9 genome and ordered them. They should arrive at the same time as the LMD9 genome. Also, as a team we decided to design primers to amplify just the Spacer/Repeat region which is around 1.4kb out of the LMD9 CRISPR genome. We noticed that there is an SpeI site present in the Spacer/Repeat region of the CRISPR and decided to find various ways to solve it.<br><br><br />
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We received our LMD9 genome from ATCC and our primers along with it. The primers have EcorI + XbaI site present going forward and SpeI + PstI sites present in the reverse primer. This was done to allow us to ligate our 1.4kb insert according iGEM biobrick standards. To solve the problem of our SpeI site at a distance of about 500bp from the leader sequence (the beginning of the Spacer/Repeat Region), we decided to use the method of Site-Directed Mutagenesis (SDM) as suggested by our advisers.<br><br><br />
<br />
We PCR amplified out the LMD 9 Crispr Spacer/Repeat region out of the genomic DNA and ran it on gel. We obtained bands at 1.4 kb, as expected.<br><br><br />
<br />
We started the process of cloning our insert in the TOPO cloning vector as the TOPO vectors are more easy to subject to SDM. For its specific protocol, refer to our protocols section. <br><br><br />
<br />
After incubation period of about 16 hours, we picked 10 colonies from the plates for mini-prepping the vector which hopefully would have our insert. The mini-prep was performed on all the ten colonies and the concentration results observed are as follows:<br><br><br />
<br />
We performed double digest on the 8 plasmids we obtained from our miniprep to verify whether the TOPO vector consist of our Spacer/Repeat Region insert. We used the EcoRI and PstI restriction sites to double digest. After obtaining the digest,we ran it on an agarose gel to verify whether the insert is present, but the results were inconclusive. We decided to continue with the SDM hoping that the insert is present.<br><br><br />
<br />
Followed the complete protocol for SDM as described on our protocols page. Tranformed and plated the bacteria and waiting for their growth.<br><br><br />
<br />
No growth observed with the SDM bacteria. We decided to revert to our backup plan of cutting our insert at the 500 bp region around the SpeI site and use that as our insert for the biobrick. Digestion and gel purification yielded 5.6 ng/ul of insert DNA.<br><br><br />
<br />
The bacterial transformation failed. Transforming biobricks again in the Novablue cells and hoping for growth.<br><br><br />
<br />
Observed colonies with pSB1C3 and innoculated them and performed miniprep on them. Digest the plasmid and do a gel purification. Ligate our insert in the plasmid vector and transform Novablue cells with it.<br><br><br />
<br />
Check for growth.... Cultures were centrifuged in the hopes that we could perform a miniprep and get DNA in time to ship it, but minimal growth was observed at 7PM. We checked the cultures again at 10:30PM and observed a significant amount of growth, but unfortunately it is too late to proceed. Hopefully next year will be better! :’(<br><br><br />
<br />
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</head><br />
</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/JulyTeam:GeorgiaTech/July2011-09-29T03:35:18Z<p>Pkurani: </p>
<hr />
<div>{{GT-header}}<br />
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<html><br />
<link href='http://fonts.googleapis.com/css?family=Josefin+Sans:400,700' rel='stylesheet' type='text/css'><br />
<head><br />
<font size="2" face="Century Gothic"><br />
<br><br><br />
Lab training sessions with our graduate TA, Sun Young Goo. The majority of the students on the team were new and had little to no lab experience. We covered all of the basics-- from simply preparing media and using an autoclave to performing genomic DNA extraction and restriction digests. We covered PCR amplification and learned the basics of primer design. Basic tutorials on bioinformatics software were provided by Kettner and Mitesh, and everyone learned how to use NEBCutter, blastn, and the art of finding the DNA sequences you are looking for on GenBank.<br><br><br />
<br />
Order Streptococcus thermophilus strain LMD-9 from ATCC, which is predicted to have an active CRISPR locus based on comparative genome sequencing done by Barrangou, et al (See citations).<br><br><br />
<br />
Learned that LMD-9 is backordered until August or September, and thus we will not be able to use it. We blasted sequences for the other two sequenced S. thermophilus strains, and chose strain LMG 13811 due to its similarity to strain LMD-9 (in respect to the repeat region). Delivery estimate for LMG is three business days. We worked on designing primers to amplify the CRISPR1/Cas region from the genomic DNA. We also discussed that while S. thermophilus LMG 18311 was predicted to have a CRISPR system, no papers had documented its functionality.<br><br><br />
<br />
We mini-prepped DNA from E. coli BL21 cells that were provided by Eric Gaucher’s lab. These cells contained a pUC18 plasmid that we would be using as our shuttling vector. The concentration of DNA from the plasmid was 200ng/uL. Using the vector map, we finalized the design of our primers with built in restriction sites to ensure that our DNA could be put in the vector and cloned in another form of E. coli (to avoid possible autoimmune problems).<br><br><br />
<br />
Still no S. thermophilus LMG 18311. We anticipate it will arrive tomorrow, and prepared 1L of LM17 media (M17 media with a lactose supplement) and 20 LM17 agar plates. We ordered primers today to amplify our CRISPR/cas locus.<br><br><br />
<br />
We have finally received our bacteria in the mail! They were sent as a lyophilized sample, which we resusitated, streaked on a agar plate, and also inoculated in liquid media (all with the appropriate antibiotics). Primers arrived in the mail and were stored in the -20C freezer.<br><br><br />
<br />
Advisor meeting today. We discussed the team project description and the safety proposal, which are due next Friday. The bacteria grown on agar were streaked on a plate, and then we performed a genomic DNA extraction on the ones that had grown in liquid media. The DNA was stored in the -20C freezer overnight.<br><br><br />
<br />
<br />
We performed a PCR of the DNA extraction product, using the primers we received on Thursday. We discussed the issue of the unknown functionality of LMG CRISPR, but hoped that we could make a major contribution to science. However, this is a huge risk to take. We decided to press on and test our CRISPR in E. coli.<br><br><br />
<br />
Today we prepared electrocompetent E. coli for transformation with our DNA + vector. We also performed a PCR clean-up of the DNA.<br><br><br />
<br />
Performed a digestion of our plasmid backbone and PCR product. We ligated these two components and transformed our E. coli. We should observe growth tomorrow.<br><br><br />
<br />
Our colonies had no growth. We had a talk with Dr. Gaucher and Dr. Hammer to figure out what to do. We are suspicious of cloning the vector in E. coli, and discussed using other organisms for our model.<br><br><br />
<br />
The team project description and the safety proposal were posted on our website. <br><br><br />
<br />
</font><br />
</head><br />
</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/JulyTeam:GeorgiaTech/July2011-09-29T03:35:04Z<p>Pkurani: </p>
<hr />
<div>{{GT-header}}<br />
{{GT-nav}}<br />
<br />
<html><br />
<link href='http://fonts.googleapis.com/css?family=Josefin+Sans:400,700' rel='stylesheet' type='text/css'><br />
<head><br />
<font size="2" face="Century Gothic"><br />
<br><br><br><br />
Lab training sessions with our graduate TA, Sun Young Goo. The majority of the students on the team were new and had little to no lab experience. We covered all of the basics-- from simply preparing media and using an autoclave to performing genomic DNA extraction and restriction digests. We covered PCR amplification and learned the basics of primer design. Basic tutorials on bioinformatics software were provided by Kettner and Mitesh, and everyone learned how to use NEBCutter, blastn, and the art of finding the DNA sequences you are looking for on GenBank.<br><br><br />
<br />
Order Streptococcus thermophilus strain LMD-9 from ATCC, which is predicted to have an active CRISPR locus based on comparative genome sequencing done by Barrangou, et al (See citations).<br><br><br />
<br />
Learned that LMD-9 is backordered until August or September, and thus we will not be able to use it. We blasted sequences for the other two sequenced S. thermophilus strains, and chose strain LMG 13811 due to its similarity to strain LMD-9 (in respect to the repeat region). Delivery estimate for LMG is three business days. We worked on designing primers to amplify the CRISPR1/Cas region from the genomic DNA. We also discussed that while S. thermophilus LMG 18311 was predicted to have a CRISPR system, no papers had documented its functionality.<br><br><br />
<br />
We mini-prepped DNA from E. coli BL21 cells that were provided by Eric Gaucher’s lab. These cells contained a pUC18 plasmid that we would be using as our shuttling vector. The concentration of DNA from the plasmid was 200ng/uL. Using the vector map, we finalized the design of our primers with built in restriction sites to ensure that our DNA could be put in the vector and cloned in another form of E. coli (to avoid possible autoimmune problems).<br><br><br />
<br />
Still no S. thermophilus LMG 18311. We anticipate it will arrive tomorrow, and prepared 1L of LM17 media (M17 media with a lactose supplement) and 20 LM17 agar plates. We ordered primers today to amplify our CRISPR/cas locus.<br><br><br />
<br />
We have finally received our bacteria in the mail! They were sent as a lyophilized sample, which we resusitated, streaked on a agar plate, and also inoculated in liquid media (all with the appropriate antibiotics). Primers arrived in the mail and were stored in the -20C freezer.<br><br><br />
<br />
Advisor meeting today. We discussed the team project description and the safety proposal, which are due next Friday. The bacteria grown on agar were streaked on a plate, and then we performed a genomic DNA extraction on the ones that had grown in liquid media. The DNA was stored in the -20C freezer overnight.<br><br><br />
<br />
<br />
We performed a PCR of the DNA extraction product, using the primers we received on Thursday. We discussed the issue of the unknown functionality of LMG CRISPR, but hoped that we could make a major contribution to science. However, this is a huge risk to take. We decided to press on and test our CRISPR in E. coli.<br><br><br />
<br />
Today we prepared electrocompetent E. coli for transformation with our DNA + vector. We also performed a PCR clean-up of the DNA.<br><br><br />
<br />
Performed a digestion of our plasmid backbone and PCR product. We ligated these two components and transformed our E. coli. We should observe growth tomorrow.<br><br><br />
<br />
Our colonies had no growth. We had a talk with Dr. Gaucher and Dr. Hammer to figure out what to do. We are suspicious of cloning the vector in E. coli, and discussed using other organisms for our model.<br><br><br />
<br />
The team project description and the safety proposal were posted on our website. <br><br><br />
<br />
</font><br />
</head><br />
</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/JulyTeam:GeorgiaTech/July2011-09-29T03:34:08Z<p>Pkurani: </p>
<hr />
<div>{{GT-header}}<br />
{{GT-nav}}<br />
<br />
<html><br />
<link href='http://fonts.googleapis.com/css?family=Josefin+Sans:400,700' rel='stylesheet' type='text/css'><br />
<head><br />
<font size="2" face="Century Gothic"><br />
<br />
Lab training sessions with our graduate TA, Sun Young Goo. The majority of the students on the team were new and had little to no lab experience. We covered all of the basics-- from simply preparing media and using an autoclave to performing genomic DNA extraction and restriction digests. We covered PCR amplification and learned the basics of primer design. Basic tutorials on bioinformatics software were provided by Kettner and Mitesh, and everyone learned how to use NEBCutter, blastn, and the art of finding the DNA sequences you are looking for on GenBank.<br><br><br />
<br />
Order Streptococcus thermophilus strain LMD-9 from ATCC, which is predicted to have an active CRISPR locus based on comparative genome sequencing done by Barrangou, et al (See citations).<br><br><br />
<br />
Learned that LMD-9 is backordered until August or September, and thus we will not be able to use it. We blasted sequences for the other two sequenced S. thermophilus strains, and chose strain LMG 13811 due to its similarity to strain LMD-9 (in respect to the repeat region). Delivery estimate for LMG is three business days. We worked on designing primers to amplify the CRISPR1/Cas region from the genomic DNA. We also discussed that while S. thermophilus LMG 18311 was predicted to have a CRISPR system, no papers had documented its functionality.<br><br><br />
<br />
We mini-prepped DNA from E. coli BL21 cells that were provided by Eric Gaucher’s lab. These cells contained a pUC18 plasmid that we would be using as our shuttling vector. The concentration of DNA from the plasmid was 200ng/uL. Using the vector map, we finalized the design of our primers with built in restriction sites to ensure that our DNA could be put in the vector and cloned in another form of E. coli (to avoid possible autoimmune problems).<br><br><br />
<br />
Still no S. thermophilus LMG 18311. We anticipate it will arrive tomorrow, and prepared 1L of LM17 media (M17 media with a lactose supplement) and 20 LM17 agar plates. We ordered primers today to amplify our CRISPR/cas locus.<br><br><br />
<br />
We have finally received our bacteria in the mail! They were sent as a lyophilized sample, which we resusitated, streaked on a agar plate, and also inoculated in liquid media (all with the appropriate antibiotics). Primers arrived in the mail and were stored in the -20C freezer.<br><br><br />
<br />
Advisor meeting today. We discussed the team project description and the safety proposal, which are due next Friday. The bacteria grown on agar were streaked on a plate, and then we performed a genomic DNA extraction on the ones that had grown in liquid media. The DNA was stored in the -20C freezer overnight.<br><br><br />
<br />
<br />
We performed a PCR of the DNA extraction product, using the primers we received on Thursday. We discussed the issue of the unknown functionality of LMG CRISPR, but hoped that we could make a major contribution to science. However, this is a huge risk to take. We decided to press on and test our CRISPR in E. coli.<br><br><br />
<br />
Today we prepared electrocompetent E. coli for transformation with our DNA + vector. We also performed a PCR clean-up of the DNA.<br><br><br />
<br />
Performed a digestion of our plasmid backbone and PCR product. We ligated these two components and transformed our E. coli. We should observe growth tomorrow.<br><br><br />
<br />
Our colonies had no growth. We had a talk with Dr. Gaucher and Dr. Hammer to figure out what to do. We are suspicious of cloning the vector in E. coli, and discussed using other organisms for our model.<br><br><br />
<br />
The team project description and the safety proposal were posted on our website. <br><br><br />
<br />
</font><br />
</head><br />
</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/FutureTeam:GeorgiaTech/Future2011-09-29T03:30:54Z<p>Pkurani: </p>
<hr />
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</center><br />
<br><br />
<br><br />
<br><br />
<br><br />
The ultimate goal of our project was to utilize this system as a mobile gene targeting system capable of eliminating antibiotic resistance. The protoype construct, pSTINGER, was designed for quantitative modeling, proof of concept, and vaccination on a single plasmid. The premise of our vaccine is that it is able to conjugate within a population and therefore an origin of transfer consistent with most conjugation mechanisms [citation] (OriT) was included. In total, the construct contains one functional gene, and that gene is Cas9. Cas9 originates from the CRISPR/Cas system of Streptococcus thermophilus LMD-9 and serves as open analog to the functionally evidenced CRISPR3 locus within commercial strain Streptococcus thermophilus DGCC7710. Cas9 requires a highly stable and characterized promoter, so a Sigma factor 70 sequence was included in the construct. Cas9 has been shown as the sole Cas gene required for CRISPR RNA directed cleaving of recognized DNA in mobile constructs also containing native CRISPR DNA[citation].<br><br><br />
<br />
In order to explicitly target DNA of interest we designed a synthetic Spacer repeat region flanked by the native leader and terminating sequence from LMD-9. Each spacer sequence of numbers 1 through 5 is a 30 base pair region with in the active functional domain of the translated genes. For functional proof of concept we included a kanamycin resistance gene to demonstrate function loss of the resistance phenotype. For quantitative modeling, we envisioned a continous flow cytometer for monitoring population containing a GFP coding target plasmid, and pSTINGER with RFP cloned in. In this experiment, we seek quantitative flourescence data to compliment our population model detailed <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Modeling">elsewhere</a>. Lastly we seek to go for the throat, we target functional domains of New Delhi metallo-beta-lactamase 1, vanA-type vancomycin resistance genes, and MecA-type methicillin resistance genes for downstream experiments.<br />
<br />
<center><br />
<img src="https://static.igem.org/mediawiki/2011/b/bd/Assembly.png" width="950" height=auto /> <br />
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</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/JulyTeam:GeorgiaTech/July2011-09-29T03:27:26Z<p>Pkurani: Created page with "{{GT-header}} {{GT-nav}} <html> <link href='http://fonts.googleapis.com/css?family=Josefin+Sans:400,700' rel='stylesheet' type='text/css'> <head> <font size="2" face="Century Go..."</p>
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</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/NotebookTeam:GeorgiaTech/Notebook2011-09-29T03:26:28Z<p>Pkurani: </p>
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<center><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/July">July Journals</a> <br><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/August">August Journals</a> <br><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/September">September Journals</a> <br />
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<br><br />
<br><br />
<center><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/July">July Journals</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/August">August Journals</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/September">September Journals</a> <br />
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<center><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/July">July</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/August">August</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/September">September</a> <br />
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</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/Week1Team:GeorgiaTech/Week12011-09-29T03:24:44Z<p>Pkurani: Blanked the page</p>
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<div></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/NotebookTeam:GeorgiaTech/Notebook2011-09-29T03:21:27Z<p>Pkurani: </p>
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<div>{{GT-header}}<br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Week1">Week 1</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week2">Week 2</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week3">Week 3</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week4">Week 4</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week5">Week 5</a> <br><br><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week6">Week 6</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week7">Week 7</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week8">Week 8</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week9">Week 9</a> <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week10">Week 10</a> <br><br><br />
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<hr />
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</center><br />
<br><br />
<br><br />
<br><br />
<br><br />
The ultimate goal of our project was to utilize this system as a mobile gene targeting system capable of eliminating antibiotic resistance. The protoype construct, pSTINGER, was designed for quantitative modeling, proof of concept, and vaccination on a single plasmid. The premise of our vaccine is that it is able to conjugate within a population and therefore an origin of transfer consistent with most conjugation mechanisms [citation] (OriT) was included. In total, the construct contains one functional gene, and that gene is Cas9. Cas9 originates from the CRISPR/Cas system of Streptococcus thermophilus LMD-9 and serves as open analog to the functionally evidenced CRISPR3 locus within commercial strain Streptococcus thermophilus DGCC7710. Cas9 requires a highly stable and characterized promoter, so a Sigma factor 70 sequence was included in the construct. Cas9 has been shown as the sole Cas gene required for CRISPR RNA directed cleaving of recognized DNA in mobile constructs also containing native CRISPR DNA[citation].<br><br><br />
<br />
In order to explicitly target DNA of interest we designed a synthetic Spacer repeat region flanked by the native leader and terminating sequence from LMD-9. Each spacer sequence of numbers 1 through 5 is a 30 base pair region with in the active functional domain of the translated genes. For functional proof of concept we included a kanamycin resistance gene to demonstrate function loss of the resistance phenotype. For quantitative modeling, we envisioned a continous flow cytometer for monitoring population containing a GFP coding target plasmid, and pSTINGER with RFP cloned in. In this experiment, we seek quantitative flourescence data to compliment our population model detailed elsewhere. Lastly we seek to go for the throat, we target functional domains of New Delhi metallo-beta-lactamase 1, vanA-type vancomycin resistance genes, and MecA-type methicillin resistance genes for downstream experiments.<br />
<br />
<center><br />
<img src="https://static.igem.org/mediawiki/2011/b/bd/Assembly.png" width="950" height=auto /> <br />
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</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/PartsTeam:GeorgiaTech/Parts2011-09-29T02:59:52Z<p>Pkurani: </p>
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[http://partsregistry.org Registry]<br />
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<groupparts>iGEM011 GeorgiaTech</groupparts></div>Pkuranihttp://2011.igem.org/File:Biobrick.pngFile:Biobrick.png2011-09-29T02:52:43Z<p>Pkurani: uploaded a new version of &quot;File:Biobrick.png&quot;</p>
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<div></div>Pkuranihttp://2011.igem.org/File:Biobrick.pngFile:Biobrick.png2011-09-29T02:51:02Z<p>Pkurani: </p>
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<a href="https://2011.igem.org/Team:GeorgiaTech">Home</a> | <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Team">Team</a> | <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Abstract">Abstract</a> |<br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Contact">Contact Us</a> |<br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/StOverview">Overview</a> |<br />
<a href="https://2011.igem.org/Team:GeorgiaTech/CRISPR">What is CRISPR?</a> |<br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Citations">Citations</a> |<br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Future">Future</a> |<br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Safety">Safety</a> | <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/SafetyTraining">Safety Training</a> | <br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Notebook">Notebook</a> | <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/StProtocols">Protocols</a> |<br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Modeling">Modeling</a> | <br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Parts">Parts Submitted to the Registry</a> | <br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Contributions">Contributions</a> |<br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Sponsors">Sponsors</a> |<br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Week1">Week 1</a> <br><br><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week2">Week 2</a> <br><br><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week3">Week 3</a> <br><br><br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Week7">Week 7</a> <br><br><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week8">Week 8</a> <br><br><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week9">Week 9</a> <br><br><br />
<a href="https://2011.igem.org/Team:GeorgiaTech/Week10">Week 10</a> <br><br><br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Week1">Week 1</a> <br><br><br />
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<groupparts>iGEM011 GeorgiaTech</groupparts></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTechTeam:GeorgiaTech2011-09-29T01:50:29Z<p>Pkurani: </p>
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Welcome to Georgia Tech iGEM 2011 Wiki!<br />
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Purpose: As a research team, our goals are two fold: to mobilize the crispr/cas bacterial immune system on a plasmid, and subsequently utilize this system as an intelligent gene targeting method capable of eliminating antibiotic resistance.<br />
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Welcome to Georgia Tech iGEM 2011 Wiki!<br />
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Purpose: As a research team, our goals are two fold: to mobilize the crispr/cas bacterial immune system on a plasmid, and subsequently utilize this system as an intelligent gene targeting method capable of eliminating antibiotic resistance.<br />
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Welcome to Georgia Tech iGEM 2011 Wiki!<br />
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Purpose: As a research team, our goals are two fold: to mobilize the crispr/cas bacterial immune system on a plasmid, and subsequently utilize this system as an intelligent gene targeting method capable of eliminating antibiotic resistance.<br />
<br />
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<br />
Purpose: As a research team, our goals are two fold: to mobilize the crispr/cas bacterial immune system on a plasmid, and subsequently utilize this system as an intelligent gene targeting method capable of eliminating antibiotic resistance.<br />
<br />
<br><br><br />
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Welcome to Georgia Tech iGEM 2011 Wiki!<br />
<br><br><br><br />
<br />
<br />
Purpose: As a research team, our goals are two fold: to mobilize the crispr/cas bacterial immune system on a plasmid, and subsequently utilize this system as an intelligent gene targeting method capable of eliminating antibiotic resistance.<br />
<br />
<br />
<br />
<a href="https://2011.igem.org/Main_Page" target="_blank"><br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Week3">Week 3</a> <br><br><br />
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<a href="https://2011.igem.org/Team:GeorgiaTech/Week9">Week 9</a> <br><br><br />
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Dr. Eric Gaucher<br />
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Dr. Eric Gaucher exploits a multidisciplinary approach towards research to generate an 'evolutionary synthetic biology'. This combination of molecular evolution and biomedicine provides a better understanding of basic molecular processes while simultaneously generating biomolecules useful for industrial and therapeutic purposes. Dr. Gaucher received his Ph.D. in 2001 and then worked for NASA until 2003 studying the Origins and Evolution of Early Life. He was then Researcher/President of a non-profit research organization until 2008 at which time he accepted an Associate Professor position at the Georgia Institute of Technology.<br />
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Dr. Brian Hammer<br />
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Dr. Brian Hammer is a molecular microbiologist interested a process of chemical communication called Quorum Sensing, which bacteria use to orchestrate group behavior. He received his MS in aquatic ecology from the University of Michigan in 1995 and his PhD in Microbiology from the University of Michigan Medical School in 2001. He was a National Science Foundation (NSF) Postdoctoral Fellow and Research Scientist at Princeton University from 2001-2008. Dr. Hammer joined Georgia Tech in 2008 where he is currently an Assistant Professor in the School of Biology. Dr. Hammer is funded by two programs at the NSF: the Division of the Molecular and Cellular Biosciences (MCB); and the Network Science and Engineering (NetSE) program. <br />
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Many of his lab members study the aquatic pathogen Vibrio cholerae, with a focus on the molecular mechanisms that permit this bacterium to convert extracellular chemical signals into changes in the expression of genes important in marine systems and the human host. In addition, other members of the Hammer lab are developing synthetic quorum sensing systems to model, simulate, and experimentally validate the fundamental limits and protocols for molecular communication. This work is in collaboration with several professors in the College of Engineering at Georgia Tech.<br />
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Dr. Harold Kim<br />
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Dr. Harold Kim is an experimental biophysicist who studies the biophysics of the genome and its influence on gene expression patterns. His research group uses single-molecule and single-cell fluorescence microscopy to understand (1)how intrinsic DNA looping dynamics is related to nucleosome formation; (2) how nucleosome positioning influences gene regulation; and (3) how spatial distribution and copy number of genes influences their expression patterns. Dr. Harold Kim received his PhD in applied physics from Stanford University in 2004, did his postdoctoral research at Harvard University from 2005 to 2009, and joined the School of Physics in 2010 as an assistant professor. He is the recipient of a Burroughs Wellcome Career Award at the Scientific Interface.<br />
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Dr. Mark Styczynski<br />
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Dr. Mark Styczynski uses experimental and computational techniques to study the connections between the different layers of regulation in cells and the cells' ultimate phenotypic outcomes. Of most interest to his lab are metabolites, the small molecule building blocks necessary for all cellular functions both as source materials and as cues that prompt regulation and other cellular responses. He studies metabolism in a variety of systems ranging from yeast to human cancer cell lines. Dr. Styczynski received his PhD from MIT in 2007, was a postdoctoral associate at the Broad Institute from 2007 to 2009, and joined the School of Chemical & Biomolecular Engineering at Georgia Tech in 2009 as an Assistant Professor.<br />
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Dr. Joshua Weitz is a quantitative biologist interested in the structure and dynamics of complex biological systems. He received his PhD in Physics from MIT in 2003 and was a NSF Postdoctoral Fellow and Associate Research Scholar at Princeton University from 2003-2006. Dr. Weitz joined Georgia Tech in 2007 where he is currently an Assistant Professor of Biology with courtesy appointments in Physics and Bioengineering. Dr. Weitz is the recipient of a Burroughs Wellcome Career Award at the Scientific Interface and is funded by the James S. McDonnell Foundation, NSF, and DARPA. His research group includes ecologists, mathematicians, physicists and bioinformaticians working on four major research themes: (i) viral dynamics at the molecular, population and evolutionary scales; (ii) quantitative systems biology and bioinformatics; (iii) structure and function of vascular networks; (iv) theoretical ecology and epidemiology. The work in the Weitz group is primarily theoretical in nature, and utilizes the tools of nonlinear dynamics, stochastic processes, and large-scale data analysis to interact with experimentalists. Examples of recent and ongoing projects include studies of collective decision making in bacterial viruses, robustness and fragility of gene regulatory networks to copy number variation, unsupervised approaches to binning short environmental sequence fragments, network phenotyping and classification of root system architecture, and a Hierarchical Bayesian analysis of allometric scaling models in biology.<br />
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Ryan Randall<br />
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"Ryan manages Prof. Gaucher's laboratory and performs original research as a 2nd year graduate student within the School of Biology at GaTech. She is currently evolving monomeric red fluorescent proteins in the laboratory. Her data will be used to construct an experimental phylogeny that will in turn be used to benchmark computational methods of ancestral sequence reconstruction. She received her BS from the University of Georgia in 2008 with a degree in Cellular Biology. She is the graduate advisor of the Georgia Tech iGEM 2011 team and has been a constant source of support for the team. She has proven invaluable to GTiGEM 2011 team and has provided us with all the lab supplies and protocols."<br />
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<br>-Mitesh Agrawal<br />
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Mitesh Agrawal<br />
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""Mitesh is a 3rd year international student from India majoring in Biomedical Engineering. Besides being all around awesome, he was a member of Georgia Tech’s first iGEM team where he was a major contributor to the modeling section of the project. This year he has once again been a vital part of the team, helping with experimental design and giving us perspective on how things worked last year. His research endeavors have included study of embryonic stem cells and genetic engineering. Mitesh was a crew member (radio specialist and biologist) for the Georgia Tech Mars Desert Research Station and he is also actively involved in the Georgia Alpha chapter of Tau Beta Pi - an engineering honors society. When not in the lab, Mitesh enjoys playing a rousing game of racquetball and reading science fiction novels. Oh, and he is a die-hard Manchester United fan, so watch out Liverpool. >:)”<br />
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Calvin Goveia<br />
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"Calvin is a 4th year chemical engineer specializing in bioprocess engineering. He is very interested in using ethanol as a fuel, and looking into the process of the utilizating of recalcitrant biomass to create renewable bioenergy. He would like to improve the current process of producing cellulose based biofuels by cloning essential enzymes from cellulolytic bacteria. He hopes to engineer E. coli with these genes to simultaneously hydrolyze cellulose and ferment those products of hydrolysis. Calvin is also working on the modeling component of our project."<br />
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<br>-Paul Sebexen<br />
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Kettner Griswold Jr.<br />
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"Kettner Griswold is a straight up mad scientist. He is a 2nd year MSE Student who hails from Bethesda, MD where he was a local legend- seriously, google him. Kettner worked at the J. Craig Venter Institue for a year during high school and now spends his days reading scientific literature and white water kayaking (usually not at the same time)."<br />
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<br>-Hannah Keith<br />
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Hannah Keith<br />
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"Hannah is a second year biochemistry major and is a vital member of the GTiGEM 2011 team. Besides being an important contributor in experimental design of this year’s project, she is the one who has kept our team organized and well run. Her field of interest is studying structural and functional properties of different biochemical molecules, including a special fondness for H2O. Hannah loves running and is an avid reader of non-fictional books. When not busy with studies and iGEM, she devotes her time to various community services and organizations like the American Medical Student Association. Overall, she has been an amazing addition to the Georgia Tech iGEM team."<br />
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<br>-Mitesh Agrawal<br />
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Priya Kurani<br />
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"Priya is a 4th year Computational Media major. She is interested in how modern media channels can help illustrate complex processes in Synthetic Biology. She likes intuitive user interfaces, Afrojack, and Cuban food."<br />
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<br>-Paul Sebexen<br />
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Paul Sebexen<br />
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"Paul Sebexen is a 3rd-year Computer Science major. He is originally from New York City and has research interests ranging from computational biology to solid-state physics. In his free time, Paul enjoys studying languages, bicycling, tennis, and playing piano. Paul loves everything German."<br />
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<br>-Priya Kurani<br />
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</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/CitationsTeam:GeorgiaTech/Citations2011-09-29T01:29:42Z<p>Pkurani: Created page with "{{GT-header}} {{GT-nav}} <html> <link href='http://fonts.googleapis.com/css?family=Josefin+Sans:400,700' rel='stylesheet' type='text/css'> <head> <font size="2" face="Century Go..."</p>
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Week 1<br />
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<center><b>Title:</b> De Novo Adaptation of Streptococcus thermophilus CRISPR1 Defense in E. coli</center><br />
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A diverse range of Bacteria and Archaea acquire resistance to foreign DNA by integrating short fragments of the invading nucleic acid into clusters of regularly interspaced short palindromic repeats (CRISPRs) on their genomic DNA. For our project we have PCR amplified the CRISPR1 locus from the chromosome of Streptococcus thermophilus [LMD-9] and ligated it into an integration vector to place it on the chromosome of Bacillus subtilis through allelic recombination on the chromosome. B. subtilis served as our model organism because it does not naturally posses a CRISPR mechanism. This should demostrate that the S. thermophilus CRISPR1/Cas system can be transferred into Bacillus subtilis and provide heterologous protection against plasmid transformation and phage infection.<br />
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<center><b>Project Overview</b><br><br></center><br />
Clusted regularly interspaced palindromic repeats (CRISPRs) and their associated cas genes confer immunity to bacteria and archaea by incorporating segments of foreign DNA, known as spacers, into their chromosome to recognize and inactivate the invader in the future. Our research project aims to clone the CRISPR1 system from Streptococcus thermophilus strain LMD-9 and transform it into Escherichia coli and Bacillus subtilis. The design of our experiment was motivated by the increasing threat of antibiotic resistant bacterial populations to human health. By designing bacteria that have a fitness advantage from the CRISPR system, we could wipe out these dangerous bacteria under specific circumstances. The cas9 gene from S. thermophilus LMD-9 works as an endonuclease, in a manner similar to RNA interference, to recognize and cleave the antibiotic resistant plasmid from populations of bacteria using predesigned spacers, such as ones for kanamycin resistance.<br />
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<center><br><br><b>Background</b><br><br></center><br />
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Streptococcus thermophilus is a lactic acid bacteria that is widely used a starter culture for yogurt products in the United States. All sequenced strains of Streptococcus thermophilus are known to have <br />
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<center><b>Project Overview</b><br><br></center><br />
Clusted regularly interspaced palindromic repeats (CRISPRs) and their associated cas genes confer immunity to bacteria and archaea by incorporating segments of foreign DNA, known as spacers, into their chromosome to recognize and inactivate the invader in the future. Our research project aims to clone the CRISPR1 system from Streptococcus thermophilus strain LMD-9 and transform it into Escherichia coli and Bacillus subtilis. The design of our experiment was motivated by the increasing threat of antibiotic resistant bacterial populations to human health. By designing bacteria that have a fitness advantage from the CRISPR system, we could wipe out these dangerous bacteria under specific circumstances. The cas9 gene from S. thermophilus LMD-9 works as an endonuclease, in a manner similar to RNA interference, to recognize and cleave the antibiotic resistant plasmid from populations of bacteria using predesigned spacers, such as ones for kanamycin resistance.<br />
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<center><br><br><b>Background</b><br><br></center><br />
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Streptococcus thermophilus is a lactic acid bacteria that is widely used a starter culture for yogurt products in the United States. All sequenced strains of Streptococcus thermophilus are known to have <br />
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</html></div>Pkuranihttp://2011.igem.org/Team:GeorgiaTech/StOverviewTeam:GeorgiaTech/StOverview2011-09-29T01:26:54Z<p>Pkurani: Created page with "{{GT-header}} {{GT-nav}} <html> <link href='http://fonts.googleapis.com/css?family=Josefin+Sans:400,700' rel='stylesheet' type='text/css'> <head> <font size="2" face="Century Go..."</p>
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<c><b>Project Overview</b><br><br></c><br />
Clusted regularly interspaced palindromic repeats (CRISPRs) and their associated cas genes confer immunity to bacteria and archaea by incorporating segments of foreign DNA, known as spacers, into their chromosome to recognize and inactivate the invader in the future. Our research project aims to clone the CRISPR1 system from Streptococcus thermophilus strain LMD-9 and transform it into Escherichia coli and Bacillus subtilis. The design of our experiment was motivated by the increasing threat of antibiotic resistant bacterial populations to human health. By designing bacteria that have a fitness advantage from the CRISPR system, we could wipe out these dangerous bacteria under specific circumstances. The cas9 gene from S. thermophilus LMD-9 works as an endonuclease, in a manner similar to RNA interference, to recognize and cleave the antibiotic resistant plasmid from populations of bacteria using predesigned spacers, such as ones for kanamycin resistance.<br />
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<c><br><br><b>Background</b><br><br></c><br />
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Streptococcus thermophilus is a lactic acid bacteria that is widely used a starter culture for yogurt products in the United States. All sequenced strains of Streptococcus thermophilus are known to have <br />
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