Mixed sequences

Bill Burnett bbur at wpo.nerc.ac.uk
Fri Jan 19 10:41:56 EST 1996


Hi All...

I'm amplifying and sequencing what I hope is bacterial 16S gene,
from a host+bacterial symbiont DNA soup...  I get very clean PCR
product using 27f and 1491r eubacterial primers, and negative 
negative controls, but mixed sequences from these products using 
the same primers for automated sequencing.  This suggests to me 
more than one species of bacteria, either a contaminant or a 
second symbiotic species or pathogen - not at all surprising. 
Just wondering if anyone else has an alternative explanation, 
or a non-cloning oriented solution. The symbiont we're aware of 
outnumbers everything else (i.e. other TEM visible bacteria) 
by at least 200:1 so we thought it'd be worth a try without cloning 
initially...  oh well...

Eternally optimistic and lazily,

Bill




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