PCR of very short DNA inserts

kiki06 · #1 (**permalink**) 03-14-2006, 10:51 PM

I have a problem. I am trying to amplify a very short DNA insert about 80 bp. The problem is my dye covers it party and also running the gel and visualizing is not easy.

Any one have any experience using primers or oligos (oligonucleotides) as inserts?

aftabac · #2 (**permalink**) 06-28-2006, 12:53 PM

hi
yes! when you have very short insert, then it is difficult to visulize it with small number of cycles, so what you can do is to increase the number of cycles of PCR reaction, so that you could get enough product that could be easily visualize.

oBWhat · #3 (**permalink**) 06-29-2006, 04:33 PM

I would run the pcr reaction on a high percentage poly acrylamide gel (like protein SDS-PAGE).

Use DNA loading dye and visualize as you would an agarose gel.
You can also use only high dye marker to prevent blocking the signal with the lower dye.

Much better!

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PCR of very short DNA inserts

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