Team:Rice University/CONSTRUCTS

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Revision as of 02:01, 30 October 2008

OUR TEAM:::SUMMARY ::: BACKGROUND ::: STRATEGY ::: CONSTRUCTS ::: RESULTS ::: ONGOING WORK

Constructs

Saccharomyces cerevisiae are widely used for baking and brewing are particularly useful for synthesizing metabolites under fermentation conditions. Such microaerobic conditions prevent the air oxidation of bioreactive compounds and are optimal for the in vivo synthesis of resveratrol. To achieve our project and expand the synthetic biology toolbox for programming yeast, we have introduced into the iGem registry BioBricks encoding 3 yeast promoters, 3 yeast terminators, a two micron origin of replication, 2 selectable markers, 2 enzymes, and a yeast integration plasmid. In addition, we have generated seven constructs using these parts. Furthermore, we have submitted two additional parts representing a foundational tool, including a gene encoding an amber suppressed RFP biobrick for screening of SupF+ (Amber suppressor) genotype and an amber suppressor tRNA biobrick.

Yeast Promoters

BBa_K122000	pPGK1	This is the 1500 bp upstream of the PGK1 coding region in an industrial yeast strain. Constitutive promoter.	1497
BBa_K122002	pADH1	700bp upstream of ADH1 promoter region containing RBS. Constitutive promoter.	701
BBa_K122017	pGAL1 + tetO	The GAL1 promoter which is highly repressed by glucose. An additional tetracycline operator site was included upstream of the RBS to allow repression by tetR.	484

Yeast Terminators

BBa_K122003	tCYC1	300bp downstream the CYC1 coding region in a standard yeast strain.	300
BBa_K122004	tADH1	300bp downstream the ADH1 coding region in a standard yeast strain.	300
BBa_K122013	tPGK1	1000bp downstream the PGK1 coding region in an industrial yeast strain.	1000

Selectable Markers

BBa_K122018	ZeoR	Zeocin Resistance Gene	300
BBa_K122008	BleoR	Bleocin Resistance Gene under pTet promoter	800ish
BBa_K122014	ORI+HisTag	2 Micron ORI and Histadine Tag	<9000

Project Specific Constructs

BBa_K122001	[pGAL1][tetO][ZeoR]	874
BBa_K122005	Tyrosine Ammonia Lyase	1933
BBa_K122010	4CL:STS	4000
BBa_K122012	[pPGK1][4CL:STS][tCYC1]	5497
BBa_K122015	[pGAL1][tetO][ZeoR][tADH1]	1175
BBa_K122021	[pADH1][TAL][tPGK1]	2651
BBa_K122019	[pPGK1][4CL:STS][tCYC1][pGAL1][tetO][ZeoR][tADH1]	1824

Additional Bacterial Parts

Novel Zero Leak Inverter (K122007 and K122006)

Amber Suppressor tRNA

K122007	The supF construct, an amber suppressor tRNA, allows for read-through at native amber (TAG) stop codons.	211
K122006	Point Mutation of RFP(13521) with incorporation of an amber stop codon at the chemophore.	923

Through incorporation of amber stop codons or point mutations of tyrosine codons (TAC) to TAG within the coding region, a genetic circuit can be used to add an additional level of regulation and determine whether a full protein or partial peptide with be synthesized. This design has characteristically high signal-to-noise ratio, with virtually no leakage.

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 ==='''Constructs'''===
-     <p>''Saccharomyces cerevisiae'' are widely used for baking and brewing, and they are particularly useful for synthesizing metabolites under fermentation conditions which prevent the air oxidation of many useful compounds. To achieve our project and expand the synthetic biology toolbox for programming yeast, we have introduced into the iGem registry BioBricks encoding 3 yeast promoters, 3 yeast terminators, a two micron origin of replication, 2 selectable markers, 2 enzymes, and a yeast integration plasmid. In addition, we have generated seven constructs using these parts.  Furthermore, we have submitted two additional parts representing a foundational tool, including a gene encoding an amber suppressed RFP biobrick for screening of SupF+ (Amber suppressor) genotype and an amber suppressor tRNA biobrick. <br />
+     <p>''Saccharomyces cerevisiae'' are widely used for baking and brewing are particularly useful for synthesizing metabolites under fermentation conditions. Such microaerobic conditions prevent the air oxidation of bioreactive compounds and are optimal for the ''in vivo'' synthesis of resveratrol. To achieve our project and expand the synthetic biology toolbox for programming yeast, we have introduced into the iGem registry BioBricks encoding 3 yeast promoters, 3 yeast terminators, a two micron origin of replication, 2 selectable markers, 2 enzymes, and a yeast integration plasmid. In addition, we have generated seven constructs using these parts.  Furthermore, we have submitted two additional parts representing a foundational tool, including a gene encoding an amber suppressed RFP biobrick for screening of SupF+ (Amber suppressor) genotype and an amber suppressor tRNA biobrick. <br />
      </p>