小柯机器人

美国麻省理工学院Bo Cao、Peter C. Dedon等研究人员合作开发出AQRNA-seq技术，实现细胞小RNA图谱的定量绘制。相关论文于2021年4月15日在线发表在《自然—生物技术》杂志上。

研究人员提出了一种方法，即绝对定量RNA测序（AQRNA-seq），该方法可最大程度地减少偏差，并为样品中所有小RNA的测序读取数和拷贝数之间提供直接的线性相关性。使用963个成员的microRNA参考文库，不同长度的寡核苷酸标准品和RNA印迹，研究人员对文库的制备和数据处理进行了优化和验证。将AQRNA-seq应用到一组人类癌细胞中，研究人员发现在癌症进展过程中可检测到超过800种可检测到的miRNA。

而在应用于细菌转移RNA库的过程中，尽管面临着二级结构和大量修饰的挑战，研究人员揭示出tRNA同受体水平的80倍变化、应激诱导的位点特异性tRNA片段化、定量修饰图谱以及应激诱导的且tRNA驱动的密码子偏向翻译。因此，AQRNA-seq提供了一种通用的方法来定量绘制细胞中的小RNA图谱。

据了解，由于文库制备过程中捕获、连接和扩增中的序列依赖性偏差，当前的下一代RNA-seq方法无法提供样品中小RNA的准确定量。

附：英文原文

Title: Quantitative mapping of the cellular small RNA landscape with AQRNA-seq

Author: Jennifer F. Hu, Daniel Yim, Duanduan Ma, Sabrina M. Huber, Nick Davis, Jo Marie Bacusmo, Sidney Vermeulen, Jieliang Zhou, Thomas J. Begley, Michael S. DeMott, Stuart S. Levine, Valrie de Crcy-Lagard, Peter C. Dedon, Bo Cao

Issue&Volume: 2021-04-15

Abstract: Current next-generation RNA-sequencing (RNA-seq) methods do not provide accurate quantification of small RNAs within a sample, due to sequence-dependent biases in capture, ligation and amplification during library preparation. We present a method, absolute quantification RNA-sequencing (AQRNA-seq), that minimizes biases and provides a direct, linear correlation between sequencing read count and copy number for all small RNAs in a sample. Library preparation and data processing were optimized and validated using a 963-member microRNA reference library, oligonucleotide standards of varying length, and RNA blots. Application of AQRNA-seq to a panel of human cancer cells revealed >800detectable miRNAs that varied during cancer progression, while application to bacterial transfer RNA pools, with the challenges of secondary structure and abundant modifications, revealed 80-fold variation in tRNA isoacceptor levels, stress-induced site-specific tRNA fragmentation, quantitative modification maps, and evidence for stress-induced, tRNA-driven, codon-biased translation. AQRNA-seq thus provides a versatile means to quantitatively map the small RNA landscape in cells.

DOI: 10.1038/s41587-021-00874-y

Source: https://www.nature.com/articles/s41587-021-00874-y

Nature Biotechnology：《自然—生物技术》，创刊于1996年。隶属于施普林格·自然出版集团，最新IF：68.164
官方网址：https://www.nature.com/nbt/
投稿链接：https://mts-nbt.nature.com/cgi-bin/main.plex