I’m going to prepare a sequencing library based on Illumina TruSeq. The protocol my experiment is based on uses single-end sequencing, but a colleague suggested that I should use the paired-end method, as it makes aligning the reads easier. Does this somehow affect the primer design and/or the indexing? I'm especially wondering if I have to use dual indexing in this case.
A colleague also suggested that I could design my own indexes in order to multiplex even more samples on the same lane when sequencing than you could while using adapters from a kit. Is it complicated to design good and functional indexes, or is it more safe to use the already published ones?
I hope these aren't completely stupid questions - and if they are, please have mercy on a poor PhD student who is just getting started with the wonderful and complex world of NGS.
Thank you already in advance for any replies, helpful links and/or suggestions!
A colleague also suggested that I could design my own indexes in order to multiplex even more samples on the same lane when sequencing than you could while using adapters from a kit. Is it complicated to design good and functional indexes, or is it more safe to use the already published ones?
I hope these aren't completely stupid questions - and if they are, please have mercy on a poor PhD student who is just getting started with the wonderful and complex world of NGS.
Thank you already in advance for any replies, helpful links and/or suggestions!
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