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bioRxiv · 10.1101/2025.11.06.686971

Hybrid untargeted and targeted RNA sequencing facilitates genotype-phenotype associations at single-cell resolution

Wang, J.·Maldifassi, M.·Bratus-Neuenschwander, A.·Zhang, Q.·Beuschlein, F.·Penton, D.·Robinson, M. D.
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Abstract

Long-read scRNA-seq enables simultaneous, unbiased identification of transcriptomic variants and gene expression but is often limited by low read coverage, restricting genotype-phenotype analyses at single-cell resolution. We systematically evaluated short-read whole-transcriptome amplification (SR-WTA) via Illumina, long-read whole-transcriptome amplification (LR-WTA), and long-read targeted sequencing (LR-Twist) via PacBio for cell typing and genotyping performance. Based on these comparisons, we propose a hybrid strategy and accompanying Snakemake pipeline that integrates SR-WTA with LR-Twist to leverage the strengths of both approaches. SR-WTA provides broad transcriptome coverage, while LR-Twist enriches a 50-gene panel for deeper variant detection. This hybrid approach improves power to link mutational profiles with transcriptional programs at single-cell resolution.

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Wang, J., Maldifassi, M., Bratus-Neuenschwander, A., Zhang, Q., Beuschlein, F., Penton, D., Robinson, M. D.. 2025-11-06. Hybrid untargeted and targeted RNA sequencing facilitates genotype-phenotype associations at single-cell resolution. https://doi.org/10.1101/2025.11.06.686971

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