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single clone isolate showed multiple bands in DGGE - (Feb/24/2006 )

Hello, everyone.

I did a clone reaction and picked up 12 isolates, extracted plasmid, PCR, run DGGE. I was expecting only one band in DGGE for each isolate, as only one DNA entered the vector. But all of the isolates gave me multiple band pattern in DGGE lane. Any idea?

Someone suggested that there may be more than one melting domains in PCR products. It sounds quite right for me. But did you ever meet such kind of gels before?




I also obtained secondary bands for for single clone isolates using DGGE. I also obtained 3 different bands for a red snow sample (18S). After excision of the bands and sequencing, they showed 3 times exactly the same sequence.

I also don't know why.

Yannick -ch

-Yannick - ch-