I'm Sanger sequencing some plasmids that are prepared from maxipreps. The sequencing results show trimmed regions (i.e. the reads are trimmed so short, because each read appears to contain multiple sequences), which cannot be used for Blast alignment analysis. I was told this is because my DNA is not from a single plasmid. Any experts can suggest what to do next?
Much appreciated.
Can we see an original .abi file of a sequence please. Sequences are usually rubbish for 30 bases after the forward primer and again on very long sequences when the background becomes comparable to the signal and this may look like 2 sequences so will be trimmed. If the whole sequence looks like 2 sequences then one possibility is that he sequencing primer has some n-1 length primer present, Another possibility is that if you are pcr ing a region of plasmid that you are not removing both primers prior to adding the sequencing primer. What method of primer removal are you using if this is the case please?
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