After I amplify the pooled sgRNA library, I need to do next-generation sequencing of the amplified sgrna library to determine sgrna distribution. So, I do the PCR as step 32 in protocol with 20ng template in 50μl reaction system with 10 reactions in total. The cycle number is 22. I got 500 μl PCR products and do PCR purif...
Im working with the Gecko v2 library and followed the Joung et al. (2017) Nature Protocols paper very closely. Now its time fpr NGS. The company which will sequence the samples asked me if there is special custom primer needed or not? The second question is: If the PCR Product is around 270 bp and you substract the adapter...
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