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1 hour ago by

Bloomington, IN

PCR amplification is common in both RNA-seq and scRNA-seq. The prevalence of duplicates in your sequenced library will be a function of library complexity (quantity of unique cDNAs before amplification) and sequencing depth. In general, the more complex the library, the deeper the library can be sequenced before you hit a wall of diminishing returns. This is because after a certain number of reads, you've sequenced most of the unique fragments in the library, and are now just sequencing PCR duplicates of those unique fragments.

RNA-seq libraries are generally complex enough that the number of sequenced PCR duplicates is negligible for analysis. scRNA-seq libraries tend to have low complexity and many PCR duplicates as a consequence, so reads are tagged with a random sequence called a unique molecular identifier (UMI) before PCR. This allows duplicate detection after sequencing since you would not really expect identical UMIs to appear in the same position/gene.

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