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  • Title: Single-cell Iso-Sequencing enables rapid genome annotation for scRNAseq analysis.
    Author: Healey HM, Bassham S, Cresko WA.
    Journal: Genetics; 2022 Mar 03; 220(3):. PubMed ID: 35143654.
    Abstract:
    Single-cell RNA sequencing is a powerful technique that continues to expand across various biological applications. However, incomplete 3'-UTR annotations can impede single-cell analysis resulting in genes that are partially or completely uncounted. Performing single-cell RNA sequencing with incomplete 3'-UTR annotations can hinder the identification of cell identities and gene expression patterns and lead to erroneous biological inferences. We demonstrate that performing single-cell isoform sequencing in tandem with single-cell RNA sequencing can rapidly improve 3'-UTR annotations. Using threespine stickleback fish (Gasterosteus aculeatus), we show that gene models resulting from a minimal embryonic single-cell isoform sequencing dataset retained 26.1% greater single-cell RNA sequencing reads than gene models from Ensembl alone. Furthermore, pooling our single-cell sequencing isoforms with a previously published adult bulk Iso-Seq dataset from stickleback, and merging the annotation with the Ensembl gene models, resulted in a marginal improvement (+0.8%) over the single-cell isoform sequencing only dataset. In addition, isoforms identified by single-cell isoform sequencing included thousands of new splicing variants. The improved gene models obtained using single-cell isoform sequencing led to successful identification of cell types and increased the reads identified of many genes in our single-cell RNA sequencing stickleback dataset. Our work illuminates single-cell isoform sequencing as a cost-effective and efficient mechanism to rapidly annotate genomes for single-cell RNA sequencing.
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