通过直接 RNA 和转录起始位点测序检测有丝分裂出芽酵母中的长未解码转录本异构体 (LUTI)。

Long undecoded transcript isoform (LUTI) detection in meiotic budding yeast by direct RNA and transcript leader sequencing.

机构信息

Department of Molecular and Cell Biology, Barker Hall, University of California, Berkeley, Berkeley, CA 94720, USA.

Department of Genome Sciences, Foege Hall, University of Washington, Seattle, WA 98105, USA.

出版信息

STAR Protoc. 2022 Feb 4;3(1):101145. doi: 10.1016/j.xpro.2022.101145. eCollection 2022 Mar 18.

DOI:10.1016/j.xpro.2022.101145

PMID:35169715

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8829799/

Abstract

LUTIs (Long Undecoded Transcript Isoforms) are 5'-extended and poorly translated mRNAs that can downregulate transcription from promoters more proximal to a gene's coding sequence (CDS). In this protocol, polyA RNA is extracted from budding yeast cells undergoing highly synchronized meiosis. Using a combination of long-read direct RNA sequencing and transcript leader sequencing (TL-seq), meiosis-specific LUTIs are systematically identified. Following identification, TL-seq is used to quantify the abundance of both LUTI and the more canonical gene-proximal (PROX) transcripts. For complete details on the use and execution of this protocol, please refer to Tresenrider et al. (2021).

摘要

LUTIs（长未解码转录本异构体）是 5'-延伸且翻译较差的 mRNAs，可下调更靠近基因编码序列（CDS）的启动子的转录。在本方案中，从经历高度同步减数分裂的出芽酵母细胞中提取 polyA RNA。使用长读直接 RNA 测序和转录本启动子测序（TL-seq）的组合，系统地鉴定减数分裂特异性 LUTIs。鉴定后，TL-seq 用于定量 LUTI 和更典型的基因近端（PROX）转录本的丰度。有关该方案使用和执行的完整详细信息，请参阅 Tresenrider 等人。（2021 年）。