IGEM:University of East Anglia (UEA), Norwich, UK/2009/Notebook/NRP-UEA-Norwich iGEM/2013/08/22: Difference between revisions
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|style="background-color: #EEE"|[[Image:igem-logo-150px.png|150px]]<span style="font-size:22px;"> Week Twelve</span> | |style="background-color: #EEE"|[[Image:igem-logo-150px.png|150px]]<span style="font-size:22px;"> Week Twelve</span> | ||
|style="background-color: #F2F2F2" align="center"| | |style="background-color: #F2F2F2" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]] }}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}} | ||
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==Floor One== | |||
Diluted soil samples 79-84 from 10^-1 to 10^-4 and plated 10^-2 to 10^-4 on SFM + Ny. | |||
Set up 38 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 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. | 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> | ||
[[Image:BIORAD 2013-08-22 14hr 32min.JPG|thumb|Fig | 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> | ||
The | 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.]] | |||
==Outreach== | |||
The registration for the jamboree was completed for the team. | |||
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Latest revision as of 23:11, 26 September 2017
Week Twelve | Main project page Previous entry Next entry |
Floor OneDiluted soil samples 79-84 from 10^-1 to 10^-4 and plated 10^-2 to 10^-4 on SFM + Ny. Set up 38 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. |