How to exchange antibiotic resistance markers on plasmids for E
(by amanosz from yahoo.com)
Tue Jun 16 08:09:18 EST 2009
Has anyone exchanged or heard of someone exchanging the antibiotic resistance marker on a plasmid for E. coli? I need to co-transform E coli with two plasmids that have the same resistance marker, and the plasmids are not available with any other marker. I have plasmid sequence information for the two plasmids of interest and a 3rd plasmid with the new resistance marker that identifies the location of each marker. I've confirmed with a BLAST search that the annotated sequences are simply the coding sequences for each marker (start to stop codon). Is it simply a modular procedure where I remove one ORF and replace it with the other? Or are there other considerations I need to think about?
The plasmid map doesn't annotate any promoter or terminator sequence for the resistance marker. But I assume if one is necessary, then I am only substituting the coding sequence and the remaining un-annotated architecture is still present.
For what it's worth, I'd like to substitute a chloramphenicol resistance marker with a kanamycin marker.
Thanks in advance for your help,
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