840:153g:Projects/project22/2012/11/15

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Prodigiosin Production in E. Coli Main project page
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PCR DNA Gel Extraction and Purification and the Beginning of T Vector Ligation

On Tuesday we ran a gel electrophoresis check on our PCR and it yielded these results:

Image:13.11.12_small.png‎ <--- Note the bands at the approximate 1500 b.p. (above the second bright line on the markers in lanes 9 and 18). Also note the functional positive control in lane 10

Based on these results, we believe we have amplified our target gene (pigI) so we extracted the DNA from the gel and purified it with a GeneJet Gel Extraction kit (Lot 00040498). We ran a gel electrophoresis check on the sample obtained from gel extraction and it yielded these results:

Image:15.11.12_small.png <-- Note the DNA at approximately 1500 b.p. (slightly above the second very bright line, which represents 1000 b.p. - slightly blurred in this picture - on the marker in lane 6)

This gel confirms our belief that we have our target gene, so we proceeded to the next step which is transformation and cloning of our DNA using a T-vector. To this end, we started the T Vector Ligation, using the Promega pGEM-T Easy Vector Ligation "Cloning PCR" protocol and will begin transformation on next Tuesday.


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