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Hi all,
Our labs protocol for metabarcoding library prep includes creating a duplicate reaction for the initial PCR step. Essentially, instead of creating a single 20uL reaction, we create two 10uL reactions in different plates. Right after running PCR, we combine the two reactions and then run a gel to make sure the PCR worked before continuing on.
The reasoning behind this as I understand it, is that we run multiple reactions to increase the odds that the PCR works to amplify our intended targets. In all my previous labs we've never split the reactions like that. It is fairly time consuming and uses more plates when we split the reaction.
My question for you all is, does anyone have experience with making duplicate reactions for library prep? Does it seem like a worthwhile thing to include in our process? We run our samples on Illumina MiSeq v3 600 cycles if that helps.
Thank you!
Angela
Our labs protocol for metabarcoding library prep includes creating a duplicate reaction for the initial PCR step. Essentially, instead of creating a single 20uL reaction, we create two 10uL reactions in different plates. Right after running PCR, we combine the two reactions and then run a gel to make sure the PCR worked before continuing on.
The reasoning behind this as I understand it, is that we run multiple reactions to increase the odds that the PCR works to amplify our intended targets. In all my previous labs we've never split the reactions like that. It is fairly time consuming and uses more plates when we split the reaction.
My question for you all is, does anyone have experience with making duplicate reactions for library prep? Does it seem like a worthwhile thing to include in our process? We run our samples on Illumina MiSeq v3 600 cycles if that helps.
Thank you!
Angela
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