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Team:Nevada/Notebook/Weeks1316
From 2011.igem.org
(→Week 13 - August 22nd-28th) |
(→Week 13 - August 22nd-28th) |
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<font color=red>E. Coli <br><br> | <font color=red>E. Coli <br><br> | ||
| - | + | <u>Bay Laurel Thioesterase</u> <br><br> | |
| + | |||
| + | MT: Five cultures of BTE coding region in the Topo vector (pUC57) were grown, and minipreped. The BTE samples and pSB1C3 were digested with Xba I and Pst I and ligated. Ligation was transformed using NEB Beta cells. Colonies were selected to culture overnight and minipreped, digested to check for correct band size. | ||
| + | |||
| + | <br> | ||
| + | |||
| + | A Free Fatty Acid Assay was done on BTE with the promoter sigma 70. A sample was also ran through a Gas chromatographer to check the length of hydrocabons we are producing. Results are in the assay section. | ||
| + | |||
| + | <br> | ||
| + | |||
| + | BTE with sigma 70 promoter and pSB1C3 were digested with EcoRI and PstI and ligated, transformed, and cultured.<br> | ||
| + | <br> | ||
| + | |||
<br> | <br> | ||
</font> <br> | </font> <br> | ||
Revision as of 19:37, 19 September 2011
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Calender Weeks 13-16
To edit on a specific week. Click on the edit button corresponding to the week.
Contents |
Week 13 - August 22nd-28th
E. Coli
Bay Laurel Thioesterase
MT: Five cultures of BTE coding region in the Topo vector (pUC57) were grown, and minipreped. The BTE samples and pSB1C3 were digested with Xba I and Pst I and ligated. Ligation was transformed using NEB Beta cells. Colonies were selected to culture overnight and minipreped, digested to check for correct band size.
A Free Fatty Acid Assay was done on BTE with the promoter sigma 70. A sample was also ran through a Gas chromatographer to check the length of hydrocabons we are producing. Results are in the assay section.
BTE with sigma 70 promoter and pSB1C3 were digested with EcoRI and PstI and ligated, transformed, and cultured.
Cyano
Comment Here
Enzymology
This week we used the GC to analyze the fatty acids being produced by one of the BTE IQ cell samples (BTE13). The fatty acids were extracted from cell supernatent using hexane and tested with a carbon disulfide solvent. We used FAME standards ranging from C:8-C:19 as well as a C:19 internal standard for comparison. Results proved positive as we were looking for C:12 FA's and there were C:8, C:10 and C:12 FA's present in the sample. We suspect that the small chains could possibly be the larger C:12 chain being metabolized and we plan to further analyzed this data. This was just a test or a pre-run and we did not use a negative control or test very many samples. We plan to do a definite run next week on four new BTE samples as well as negative controls of wild-type IQ cells and a known concentration of a C:17 internal standard will be used for quantification. Also this week we performed our third ethanol assay using the BioSystems Ethanol assay kit on 4 of the ADH/PDC IQ cell cultures. We retrieved positive results for one of the samples (ACH/PDC17) with an ethanol production of 3.4mM. We are planning to perform the ADH activity assay on this culture and four new samples next week.
Media
Comment Here
Week 14 - August 29th- September 4th
E. Coli
Comment Here
Cyano
Comment Here
Enzymology
Comment Here
Media
Comment Here
Week 15 - September 5th-11th
E. Coli
Comment Here
Cyano
Comment Here
Enzymology
Comment Here
Media
Comment Here
Week 16 - September 12th-18th
E. Coli
Comment Here
Cyano
Comment Here
Enzymology
Comment Here
Media
Comment Here
Week 17 - September 19th-25th
E. Coli
Comment Here
Cyano
Comment Here
Enzymology
Comment Here
Media
Comment Here
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