PCR ghost bands

Emma Macfarlane emma at HERA.MED.UTORONTO.CA
Mon Jun 27 10:11:07 EST 1994


Pierre, with regard to your comment that there are no obvious internal 
priming sites in your sequence, a summer student in our lab recently 
tried to amplify a gene from a plasmid construct sent to us by another
lab.  She got a band of the correct size yet when we sub-cloned and 
sequenced it we found that something strange had happened to the 3' end
To cut a long story short we discovered that the original construct had
been missing the 3' end of the gene and consequently the primer site was
non-existent - but we got a lovely strong band from the PCR!  When I 
checked the sequence for an 'obvious internal priming site' the best I 
could come up with was 3 complementary bases, a gap of four bases then 3 
more complementary bases for a primer that was 24 bases in length.  PCR
moves in mysterious ways!
Emma



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