Transition from a prokaryotic bacteria to yeast.

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1. Subcloning into a yeast expression plasmid.
1. Subcloning into a yeast expression plasmid.
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Lucas Argueso claims that this is quite easy.
2. Codon bias.
2. Codon bias.
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I'm still unsure of the process and necessity, but we may need to use different codons in Yeast than are used in E. Coli.  So, I've tried to find some info on the codons in both. [http://www.jbc.org/content/257/6/3026.full.pdf Yeast Codon bias]. [http://www.sci.sdsu.edu/~smaloy/MicrobialGenetics/topics/in-vitro-genetics/codon-usage.html E. Coli Codon bias]. [http://mbe.library.arizona.edu/data/1985/0201/2ikem.pdf Direct Comparison of E.coli and Yeast].
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The issue in Codon bias is that different organisms can produce different RNA's much easier.  If the RNA used in a codon is uncommon, it is likely to die off as was shown in the following Stanford research paper:
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[http://www.annualreviews.org/doi/pdf/10.1146/annurev.genet.42.110807.091442 Codon bias]. Using the codon bias tools below, there is a strong indication that the Bio-brick UW developed last year uses a lot of RNA uncommon in yeast. It is likely that this strain will die out in yeast very quickly. However, the Codons can be altered to use more common RNA, which would make the odds of replication greater. An optimized sequence is given by the final two links, and I am now investigating methods of synthesizing such a sequence.
[http://www.genscript.com/cgi-bin/tools/rare_codon_analysis This is a tool for analyzing the codon bias of a given protein strand in a specific organism].
[http://www.genscript.com/cgi-bin/tools/rare_codon_analysis This is a tool for analyzing the codon bias of a given protein strand in a specific organism].
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[http://www.vivo.colostate.edu/molkit/translate/index.html This gives optimized nucleic acid sequence for a given amino acid sequence].
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[http://www.jcat.de/ Another tool for analyzing the codon bias of a given protein strand in a specific organism].
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[http://www.encorbio.com/protocols/Codon.htm This gives optimized nucleic acid sequence for a given amino acid sequence].
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3. Gene synthesis.
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Chemical synthesis is done by chemically creating the desired sequence.  There is a lab on campus which would do this for us if we desired so: [http://www.pmf.colostate.edu/genomics_pricing.html Chemical Synthesis at CSU].
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4. Secreting enzyme into beer.
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3. Secreting enzyme into beer.
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Guy has done quite a bit of research on secretion/expression vectors. See his page for more information, but a useful background article is given below.
https://static.igem.org/mediawiki/2008/2/27/JHU_0708_paper_ForeignGeneExpression.pdf
https://static.igem.org/mediawiki/2008/2/27/JHU_0708_paper_ForeignGeneExpression.pdf

Latest revision as of 22:56, 27 June 2012

Home Team Official Team Profile Project Parts Submitted to the Registry Modeling Notebook Safety Sponsors Attributions

Currently I'm looking into 3 aspects of the transition to yeast:

1. Subcloning into a yeast expression plasmid.

Lucas Argueso claims that this is quite easy.

2. Codon bias.

The issue in Codon bias is that different organisms can produce different RNA's much easier. If the RNA used in a codon is uncommon, it is likely to die off as was shown in the following Stanford research paper: Codon bias. Using the codon bias tools below, there is a strong indication that the Bio-brick UW developed last year uses a lot of RNA uncommon in yeast. It is likely that this strain will die out in yeast very quickly. However, the Codons can be altered to use more common RNA, which would make the odds of replication greater. An optimized sequence is given by the final two links, and I am now investigating methods of synthesizing such a sequence.

This is a tool for analyzing the codon bias of a given protein strand in a specific organism.

Another tool for analyzing the codon bias of a given protein strand in a specific organism.

This gives optimized nucleic acid sequence for a given amino acid sequence.

3. Gene synthesis. Chemical synthesis is done by chemically creating the desired sequence. There is a lab on campus which would do this for us if we desired so: Chemical Synthesis at CSU.

4. Secreting enzyme into beer.

Guy has done quite a bit of research on secretion/expression vectors. See his page for more information, but a useful background article is given below.

https://static.igem.org/mediawiki/2008/2/27/JHU_0708_paper_ForeignGeneExpression.pdf

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