What are the polymerases that can utilised for bisulfite converted DNA, I am getting smear while doing PCR for the samples.
Most polymerases will work. A picture of the amplified product would help but over amplification may be to blame. Is your" no template control" a clean negative and how much dna are you amplifying in each reaction and for how many cycles of pcr
I am doing Bisulfite conversion of the DNA extracted from the tissues. The initial amount of DNA taken for bisulfite conversion is 150-200ng. After conversion, I am getting around 100ng of bisulfite-converted DNA (Elution in 10ul volume). For a different region, I am getting my desired band, but for another gene region, I am not getting bands that I was getting for the same PCR composition. Usually I take around 2ul (15~20ng) of template BS DNA and primer concentration of .4uM, 2.5mM MgCl2 final in 25ul PCR reaction
A smear can result frpm over amplification (often from contamination) so a ntc sample is needed. The reagents look ok so another reason for a smear is that only one primer is annealing and you are seeing the result of single primer amplification. If one primer is not annealing it may be either that it cannot find good homology or the annealing temp is too high for one primer.Try a lower Ta
KAPA HiFi HotStart Uracil+ ReadyMix PCR Kit from Roche
https://rochesequencingstore.com/wp-content/uploads/2017/10/KAPA-HiFi-HotStart-Uracil-ReadyMix-TDS_KR0413_v4.17-1.pdf
https://sequencing.roche.com/us/en/products/group/kapa-hifi-uracil-kit.html
PfuTurbo DNA Polymerase from Agilent
https://www.agilent.com/en/product/polymerase-chain-reaction-(pcr)/pcr-enzymes-reagents/high-fidelity-dna-polymerases-for-pcr/pfuturbo-dna-polymerase-785912
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