PCR product dimer - clean it up (Dec/28/2008 )
Because we are testing tyhe expression pattern of alternative splicing variant, we usually got multi-bands from single primer set.
However, when we sequencing PCR product directly each band actually is the hybride of several bands, and I was told in this Forum that it is "product dimer".
I understand cloning is the best way to do for this kind of problem, but my boss not willing to take $$$ and time using TA cloning.
Is there any way I can get reduce "product dimer" so I can have a clear sequencing result?
-wuxx0153-
you can gel purify your PCR product and extract your desired band.
-perneseblue-
QUOTE (wuxx0153 @ Dec 29 2008, 06:56 AM)
Because we are testing tyhe expression pattern of alternative splicing variant, we usually got multi-bands from single primer set.
However, when we sequencing PCR product directly each band actually is the hybride of several bands, and I was told in this Forum that it is "product dimer".
I understand cloning is the best way to do for this kind of problem, but my boss not willing to take $$$ and time using TA cloning.
Is there any way I can get reduce "product dimer" so I can have a clear sequencing result?
However, when we sequencing PCR product directly each band actually is the hybride of several bands, and I was told in this Forum that it is "product dimer".
I understand cloning is the best way to do for this kind of problem, but my boss not willing to take $$$ and time using TA cloning.
Is there any way I can get reduce "product dimer" so I can have a clear sequencing result?
I probably have met the same question as you.But the AT conten is high as 60%.The efficiency of my second PCR is very slow.I think it`s resulted by the complicated secondary structure of my target sequence also.I have used the PCR buffer for complicated template and no ideal result.
-ZGJABCXYZ-