Hello all,
This may be a naive question, but has anyone encountered a PCR product migrating to lower molecular weight on a 1.5% agarose gel (~1/2 the expected MW). Happened to 2 PCRs with different amplicons and primers sets. Ladder migrates accordingly with the loading dye.
Thanks for any potential insight!
Hi Dennis,
It is not naive question. Some that effect happens commonly when the loading and running buffer have different conductivity.
Do you use a commercial loading buffer or you make your own?
Hi Mario,
We cast our own gels and make running buffer ourselves. Everything is made in 1x TEA buffer. Samples are PCR so are stored in PCR water and the ladder is straight out of the freezer (whatever Invitrogen makes it in). Hope this makes sense.
Dennis.
Is the band very thick? If so, they can run distorted, usually at an apparent lower MW.
Try to dilute the ladder with 1X PCR buffer, it might be the salt in the PCR buffer that makes migrating the DNA faster, with this trick you are running ladder and amplicons at same salt concentration and same speed.
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