Odd band size with TexasRed-dATP PCR

C Coward cc122 at mole.bio.cam.ac.uk
Thu Mar 30 09:59:51 EST 2000


Greetings.

I am attempting to label DNA with TexasRed by incorporating TexasRed-dATP
in a PCR reaction. The expected product size is 2kb. In a control reaction
(unlabelled dATP) I generate a product of the expected size (by agarose
electrophoresis). With the TR-dATP I see a large fluorescences signal at
approx. 1.25 kb (with a fainter smear above this to approx. 2kb). I assume
(though haven't checked) that unincorporated TR-dATP would run very fast
(at least not up in the 1-2kb range). Any explanations for the
observation?

I'm wondering whether TR incorporation would change the mobility of the
DNA fragment (though if anything I would expect the mobility to be lower
due to incorporation of the large TR moiety. Alternatively if the TR
imparts a different charge to the DNA it could run faster).

Thanks in advance for any suggestions.

Chris





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