Hi!
I have ordered the Illumina PE primers and PE adapters from IDT with the required modifications. I have combed through the forum, but have not been able to find answers to two of my questions. There are references to method sections from published papers, nevertheless, I would like to know what people are currently doing for the following:
1) What solution do you re-suspend the (i) adapters and (ii) primers in?
2) What protocol do you use for annealing the adapters? Specifically, what concentration are the adapters when you heat to 95 degrees? (Is it at the stock concentration of 100 uM or the final concentration of 15 uM?) Do you let the adapters cool down to RT in the PCR machine, or just take it out of the heat-block and leave it on the bench? Is there a way of confirming that the adapters annealed properly?
I would truly appreciate hearing back. Thank you all VERY much!
Regards,
-Sushma
I have ordered the Illumina PE primers and PE adapters from IDT with the required modifications. I have combed through the forum, but have not been able to find answers to two of my questions. There are references to method sections from published papers, nevertheless, I would like to know what people are currently doing for the following:
1) What solution do you re-suspend the (i) adapters and (ii) primers in?
2) What protocol do you use for annealing the adapters? Specifically, what concentration are the adapters when you heat to 95 degrees? (Is it at the stock concentration of 100 uM or the final concentration of 15 uM?) Do you let the adapters cool down to RT in the PCR machine, or just take it out of the heat-block and leave it on the bench? Is there a way of confirming that the adapters annealed properly?
I would truly appreciate hearing back. Thank you all VERY much!
Regards,
-Sushma
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