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4

I'm not completely clear when you say "what makes the replication terminate when the polymerase reaches the primer at the other end" since when you perform a PCR you go through three phases. The denaturation, whereby the two DNA strands become single stranded, then the annealing, which is when primers attach to their appropriate matching site (but the ...


7

Note: In your PCR program you always set extension time. Case: Product length = 500bp PCR extension time = 50sec Assuming that polymerase adds 1000 nt/min Cycle 1: Strand that binds FP: extends ~800nt to the right (as per the polymerization rate): 300 bp ahead of RP complementary site. This product is lets say P1 Strand that binds RP: extends ~800nt ...


1

First as you mentioned, which is I think the key thing, is that you need to clarify what DNA sample it is that you are observing in the gel. The best thing to do to ensure that you only have PCR generated DNA samples is that once your PCR is over, treat your DNA mixture with Dpn I enzyme, which cuts methylated DNA, which is essentially cellular DNA as they ...


1

After you check your gel it would behoove you to check primers, preparation, the quality of the XNA itself, then proper electrophoresis levels and other mechanical failures. I would think improper resistance of the electrophoresis wiring would cause your problem.


5

There is one simple reason for that, your agarose gel is most likely too dense. Depending in the type of agarose, I would prepare a 0.5-0.6% gel at maximum. Synbio gives this list for "standard" agarose, which fits pretty good with my experiences. If you use low melting agarose, this table looks a bit different, as the gel matrix is not a dense. The ...



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