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. 2022 Jun 7;25(7):104530. doi: 10.1016/j.isci.2022.104530

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© 2022 The Authors

This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

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Library prepration and sequencing templates for short- and long- reads

(A) Overview of the library preparation: Microfluidic chips are used to generate 10x Chromium GEMs, which tag the RNA transcripts with cellular barcodes. After the transcripts are tagged, the GEMs are burst, and the tagged RNA material is split into two pools of sequencing, one for SRs and one for LRs.

(B) The SR template. Inside the GEMs, RNA transcripts are tagged with an Illumina adapter that is a fixed sequence, followed by a 16bp cellular barcode (CB), followed by a random 10bp sequence for the unique molecular identifier (UMI).

(C) The LR template. Note that the LR template is essentially the same as the SR template with the LR sequencing adapter added. Depending on the specifics of the library preparation, LRs may sequence the forward or the reverse strand of the RNA molecule. In either case, we expect the cellular barcode to be adjacent to the SR adapter sequence.