AI Article Synopsis
- The research focuses on the significance of polyadenylation in the stabilization and translation of messenger RNAs, leading to the development of a new method called SM-PATseq for analyzing poly(A) tail lengths in the entire transcriptome.
- SM-PATseq uses a specific cDNA synthesis technique involving oligo-dT and random hexamer priming, allowing for precise measurement of tail lengths and detection of non-A bases.
- The method is designed for use with long-read sequencing technologies, such as the Pacific Biosciences Sequel platform, and aims to improve the quantification of transcript abundance similar to traditional RNA sequencing approaches.
Article Abstract
The polyadenylation of the 3' ends of messenger RNAs is an important regulator of stability and translation. We developed the single-molecule poly(A) tail sequencing method, SM-PATseq, to assay tail lengths of the whole transcriptome at nucleotide resolution using long-read sequencing. This method generates cDNA using an oligo-dT 3' splint adaptor ligation to prime first-strand cDNA synthesis, followed by random hexamer priming for second-strand synthesis. By directly sequencing the cDNA on long-read platforms, we can resolve tail lengths at nucleotide resolution, identify non-A bases within the tail, and quantify transcript abundance analogous to traditional RNAseq methods. Here, we discuss the method for generating, sequencing, and primary analysis of poly(A) tail data from total RNA using the Pacific Biosciences Sequel platform.
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| http://dx.doi.org/10.1007/978-1-0716-3481-3_17 | DOI Listing |
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