Trouble with PCR on genomic tomato DNA (I've tried many fixes)
Posted 23 July 2009 - 11:52 AM
I have tried adding DMSO, Betaine, and BSA, with no improvement to the product. I have also tried using different reagents, I have had new primers made, I have tried hot start reactions. I have tried adjusting the ammount of MgCl2, KCl, Taq, and dNTPs. I have used a gene clean turbo kit to attempt to remove any polyphenols etc that could be inhibiting PCR.
I keep getting these short smears when I run gels. I have used control DNA and control primers to ensure that the thermocycler works propperly, once yielding exactly the product it was supposed to and more recently giving me the expected product in addition to smears that are the same as I am getting when I try to use the tomato DNA and tomato primers.
This is work that has been published in many papers with no problems. I am following their protocols exactly. Any suggestions to what might be going on with my attempt to amplify my gene of interest? Thanks
Posted 23 July 2009 - 07:54 PM
Posted 24 July 2009 - 07:53 PM
Posted 29 July 2009 - 11:43 AM
Posted 29 July 2009 - 06:33 PM
Posted 29 July 2009 - 06:54 PM