Again, I made an electrophoresis gel to see if I correctly extracted the plasmids pBBRMCS5 and pSB4C5 that were isolated last week.
I used the last 5 microliters (there is no more plasmid) and followed the same procotol as the other gels I've run. The only difference is that I set the electrophoresis chamber to 100 volts and to 70 minutes. Approximately at 6:15 p.m. I was about to stain the gel with ethidium bromide when some labmates told me the reason I couldn't clearly see something in the last gels was that I hadn't left the extractions to be digested to linearize the plasmids. The fuzzy and continuous banding observed is due to the several isoforms (concatamers) that exist in plasmidic DNA.
Nonetheless, this is the gel:
Next time, I surely will not forget to digest the extracted plasmids. I left incubating more DH5α E. coli carrying the plasmids in liquid medium overnight. 4 test tubes were used per each plasmid, with 5 ml of liquid medium. 3 test tubes (for each plasmid) were added with the respective antibiotic (5 microliters of chroramphenicol and 5 microliters of gentamicin, for pSB4C5 and pBBRMCS5, respectevely). The 4th test tubes were added with the proper bacteria without the antibiotic.
UNAM Genomics Mexico 2011
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