Team:LMU-Munich/Germination Stop

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Based on the work of [http://www.ncbi.nlm.nih.gov/pubmed/19554258 J. Kim and W. Schumann (2009)] and [http://www.ncbi.nlm.nih.gov/pubmed/11466293 B. Setlow et al (2001)], we decided to knock out genes ''cwlJ'', ''sleB'', ''cwlB'', ''gerD'', and ''cwlD''. The genes ''cwlJ'' and ''sleB'' code for lytic enzymes which are active in the process of germination. In the work of [http://www.ncbi.nlm.nih.gov/pubmed/11466293 B. Setlow et al (2001)], when ''cwlJ'' and ''sleB'' were knocked out together, germination frequency was reduced by 5 orders of magnitude. [http://www.ncbi.nlm.nih.gov/pubmed/19554258 J. Kim and W. Schumann (2009)] report a similar reduction in germination when ''gerD'' and ''cwlB'' are knocked out together, and a reduction in germination ability when the gene ''cwlD'' is knocked out. When all five of these genes are knocked out, we hope to yield a ''B. subtilis'' strain which produces spores completely incapable of germination.  
Based on the work of [http://www.ncbi.nlm.nih.gov/pubmed/19554258 J. Kim and W. Schumann (2009)] and [http://www.ncbi.nlm.nih.gov/pubmed/11466293 B. Setlow et al (2001)], we decided to knock out genes ''cwlJ'', ''sleB'', ''cwlB'', ''gerD'', and ''cwlD''. The genes ''cwlJ'' and ''sleB'' code for lytic enzymes which are active in the process of germination. In the work of [http://www.ncbi.nlm.nih.gov/pubmed/11466293 B. Setlow et al (2001)], when ''cwlJ'' and ''sleB'' were knocked out together, germination frequency was reduced by 5 orders of magnitude. [http://www.ncbi.nlm.nih.gov/pubmed/19554258 J. Kim and W. Schumann (2009)] report a similar reduction in germination when ''gerD'' and ''cwlB'' are knocked out together, and a reduction in germination ability when the gene ''cwlD'' is knocked out. When all five of these genes are knocked out, we hope to yield a ''B. subtilis'' strain which produces spores completely incapable of germination.  
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[[File:germination_gene_knockouts.jpg|650px]]
 
Two methods are being employed to knock out germination: resistance cassette (RC) knockouts and clean deletions. Single RC knockouts were created first; then they were combined to create multiple knockouts.
Two methods are being employed to knock out germination: resistance cassette (RC) knockouts and clean deletions. Single RC knockouts were created first; then they were combined to create multiple knockouts.
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[[File:germination_gene_knockouts_resized.jpg|frame|center|Fig. 1: The four germination genes being knocked out, and their resistance cassette replacements, as shown on the ''Bacillus'' chromosome.]]
This will be checked with germination assays.  
This will be checked with germination assays.  

Revision as of 08:32, 12 September 2012

iGEM Ludwig-Maximilians-Universität München Beadzillus

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The LMU-Munich team is exuberantly happy about the great success at the World Championship Jamboree in Boston. Our project Beadzillus finished 4th and won the prize for the "Best Wiki" (with Slovenia) and "Best New Application Project".

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