Hi.
I would like to use my own sequencing primer. I design it with properties (Tm
and GC content and length etc) close to standard Illumina Sequencing Primer.
To check its performance I set up a PCR reaction with the primer as forward and P7 primer as reverse. But I don't know what is the annealing temperature I should use.
Anyone know what is the temperature for Illumina Sequencing Primer to anneal during cBOT last step (not sure if it's correct)? So that I can follow the temperature in my PCR step to see if my primer can anneal.
Please help. Thanks
- NGS newbie.
I would like to use my own sequencing primer. I design it with properties (Tm
and GC content and length etc) close to standard Illumina Sequencing Primer.
To check its performance I set up a PCR reaction with the primer as forward and P7 primer as reverse. But I don't know what is the annealing temperature I should use.
Anyone know what is the temperature for Illumina Sequencing Primer to anneal during cBOT last step (not sure if it's correct)? So that I can follow the temperature in my PCR step to see if my primer can anneal.
Please help. Thanks
- NGS newbie.
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