IGEM:University of East Anglia (UEA), Norwich, UK/2009/Notebook/NRP-UEA-Norwich iGEM/2013/08/22: Difference between revisions
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=Floor One== | ==Floor One== | ||
Diluted soil samples 79-84 from 10^-1 to 10^-4 and plated 10^-2 to 10^-4 on SFM + Ny. | Diluted soil samples 79-84 from 10^-1 to 10^-4 and plated 10^-2 to 10^-4 on SFM + Ny. | ||
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==Floor Two== | ==Floor Two== | ||
We discovered after analysing the sequencing data from 18/07 that sample 3a had the correct sequence for biobrick BBa_K0141000 but sample 3b did not. This explained why sample 3b2, which was a miniprep from 3b, had extra NdeI sites. To confirm our theory, samples 3a, 3b, 3c and the original BBa_J04450 were cut with restriction enzyme pvuII and analysed by gel electrophoresis ''fig 3''. | We discovered after analysing the sequencing data from 18/07 that sample 3a had the correct sequence for biobrick BBa_K0141000 but sample 3b did not. This explained why sample 3b2, which was a miniprep from 3b, had extra NdeI sites. To confirm our theory, samples 3a, 3b, 3c and the original BBa_J04450 were cut with restriction enzyme pvuII and analysed by gel electrophoresis ''fig 3''. <br> | ||
The results show that sample 3b does appear to have extra DNA as the lowest band is largest than the ones for 3b and cut BBa_J04450. So the excised gel from the previous day was ignored. <br> | |||
Although colonies were visible on the LB plates (BL21 cells containing pETAntA and antGpRFP plasmids) they do not appear red. The plates were left to incubate longer at 37°C and the cells were harvested from the two cultures. | |||
[[Image:BIORAD 2013-08-22 14hr 32min.JPG|thumb|Fig 3:Analysis of Restriction enzyme digest with PvuII. Lanes 2 and 3 contain uncut BBa_J04450 and BBa_J04450 digested with PvuII respectively. Lanes 5-7 contain samples 3a,3b and 3c digested with PvuII respectively.]] | [[Image:BIORAD 2013-08-22 14hr 32min.JPG|thumb|Fig 3:Analysis of Restriction enzyme digest with PvuII. Lanes 2 and 3 contain uncut BBa_J04450 and BBa_J04450 digested with PvuII respectively. Lanes 5-7 contain samples 3a,3b and 3c digested with PvuII respectively.]] | ||
==Outreach== | ==Outreach== | ||
Revision as of 06:05, 23 August 2013
 Week Twelve
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Floor OneDiluted soil samples 79-84 from 10^-1 to 10^-4 and plated 10^-2 to 10^-4 on SFM + Ny. Set up more Bioassays from spore stocks - spotted 2 x 5 ul of spore stock onto a single SFM plate. The plates were left to grow at 30 ° for 7 days. Set up overnight ETpuz cultures for tomorrow's conjugation. Added 50 ul E. coli cells to ~ 10 ml LB (lacking NaCl for ETpuz pMS82 cells as Hygromycin is salt-intolerant). Added 10 ul Chloramphenicol and Kanamycin to both ETpuz pAU3-45 and pMS82 cultures, and 10 ul of Apr for pAU3-45 and 10 ul of Hyg for pMS82. Floor TwoWe discovered after analysing the sequencing data from 18/07 that sample 3a had the correct sequence for biobrick BBa_K0141000 but sample 3b did not. This explained why sample 3b2, which was a miniprep from 3b, had extra NdeI sites. To confirm our theory, samples 3a, 3b, 3c and the original BBa_J04450 were cut with restriction enzyme pvuII and analysed by gel electrophoresis fig 3.  OutreachThe registration for the jamboree was completed for the team. | |
