840:153g:Projects/project25/2012/11/29: Difference between revisions
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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Cloning of mreB from ''Caulobacter crescentus'' into ''E.Coli''</span> | |style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Cloning of mreB from ''Caulobacter crescentus'' into ''E.Coli''</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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None of our plasmids seemed to have our mreB gene insert. We were expecting two bands; 1200bp and 2700bp, our insert+promoter and plasmid backbone. It looks like we got our plasmid back bone and our promoter without the gene insert (100-200bp). Well 1 had 3 bands, which suggests that it was not fully digested. The 3 bands represent super coiled, relaxed and linear DNA; while the other samples had Super coiled and linear DNA. We decided to send 2 samples (L1A and L3B) to be sequenced as a second verification method. | None of our plasmids seemed to have our mreB gene insert. We were expecting two bands; 1200bp and 2700bp, our insert+promoter and plasmid backbone. It looks like we got our plasmid back bone and our promoter without the gene insert (100-200bp). Well 1 had 3 bands, which suggests that it was not fully digested. The 3 bands represent super coiled, relaxed and linear DNA; while the other samples had Super coiled and linear DNA. We decided to send 2 samples (L1A and L3B) to be sequenced as a second verification method. | ||
'''Thursday''': | '''Thursday''': | ||
Latest revision as of 22:18, 26 September 2017
 Cloning of mreB from Caulobacter crescentus into E.Coli
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 Main project page Previous entry
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Experiment 8: Gel Electrophoresis Verification and Semester Wrap UpTuesday: We digested our 12 miniprep samples with EcoR1 and Pst1. Then we ran out the samples on a gel to determine if our plasmids had the correct part inserted.
Well 1-4: samples L3D-L3A. Well 5: 100bp DNA ladder Plus. Well 6-9: samples L2D-L2A. Well 10: 100bp DNA Ladder Plus. Wells 11-14: samples L1D-L1A. None of our plasmids seemed to have our mreB gene insert. We were expecting two bands; 1200bp and 2700bp, our insert+promoter and plasmid backbone. It looks like we got our plasmid back bone and our promoter without the gene insert (100-200bp). Well 1 had 3 bands, which suggests that it was not fully digested. The 3 bands represent super coiled, relaxed and linear DNA; while the other samples had Super coiled and linear DNA. We decided to send 2 samples (L1A and L3B) to be sequenced as a second verification method.
We grew 10 overnight colonies, 4 from plate L1 and 6 from plate L3, to create glycerol stocks for future use. These are 10 new colonies, so we can hopefully get our gene insert in the plasmid.
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