After I obtained the results from sequencing, I proceeded to curate the sequences and identifying (with Vector NTI) the primers I used in the sequence. The primers, although are degenerated, were found with no mismatches.
The problem is that when I blast the squence using BLASTn or BLASTX, the results are malate synthase, a gene/protein that I totally didn't expected and totally unrelated to the gene I'm interesed. These results were like 13 of the 20 sequences while 5 were of the gene I was expecting.
Please tell me what went wrong. I'm a bit desperate. I used a clone library with pGEM and chemocompetent DH5-alpha.
BLAST is giving me a result totally unrelated to the gene I expect
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