Priming

... length of primer in PCR. But please if you explain the success of RAPD-PCR which use oligo decamere nucleotides as primer and there were no misspriming or false priming. Q2: If there is a software to design RAPD primer? sam fisher

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... 10 nucleotide) primers of arbitrary sequences. Polymorphism of amplified fragments are caused by: (1) base substitutions or deletions in the priming sites, (2) Insertions that render priming sites too distant to support amplification, or (3) insertions or deletions that change the size of ...

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... Sorry. That stuff about primers was because I assumed you were working with a cDNA library already cloned into plasmids. The comments about mis-priming etc don't apply at this stage.

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... or – Mg would be better; you’ll just have to try both and see. You could Blast your primer sequences to see if you might have a problem with false priming either within the gene itself, or in other genes. You could also try KOD polymerase instead of the mixture in the Expand system. (KOD prefers ...

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... can check oligos for self-loop-stem structures at http://www.basic.northwestern.edu/biotools/oligocalc.html As for checking for potential false priming, there isn’t any terribly good program I know of. I usually Blast the oligos against the target sequence at low stringency and look to see if ...

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