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A novel strand-specific RNA-sequencing protocol using dU-adaptor-assembled Tn5

文献类型： 外文期刊

作者： Tao, Xiaoyuan 1 ; Feng, Shouli 2 ; Li, Sujuan 1 ; Chen, Guang 1 ; Wang, Jian 1 ; Xu, Lizhi 1 ; Fu, Xujun 3 ; Yu, Jing 4 ; Xu, Shengchun 1 ;

作者机构： 1.Zhejiang Acad Agr Sci, Cent Lab, State Key Lab Managing Biot & Chem Threats Qual &, Hangzhou 310021, Peoples R China

2.Zhejiang Univ, Coll Agr & Biotechnol, Hangzhou 310058, Peoples R China

3.Zhejiang Acad Agr Sci, Inst Crop & Nucl Technol Utilizat, Hangzhou 310021, Peoples R China

4.Zhejiang Future Gene Biotechnol Co, Jingning 323500, Peoples R China

关键词： dU-Tn5; dUTP; lncRNA; soybean; stranded RNA-seq; Tn5

期刊名称：JOURNAL OF EXPERIMENTAL BOTANY （ 影响因子：6.9； 五年影响因子：8.0 ）

ISSN： 0022-0957

年卷期： 2023 年 74 卷 6 期

页码：

收录情况： SCI

摘要： Strand-specific RNA-seq is a powerful tool for the discovery of novel transcripts, annotation of genomes, and profiling of gene expression levels. Tn5 transposase has been successfully applied in massive-scale sequencing projects; in particular, Tn5 adaptor modification is used in epigenetics, genomic structure, and chromatin visualization. We developed a novel dU-adaptor-assembled Tn5-mediated strand-specific RNA-sequencing protocol and compared this method with the leading dUTP method in terms of experimental procedure and multiple quality metrics of the generated libraries. The results showed that the dU-Tn5 method is easy to operate and generates a strand-specific RNA-seq library of comparable quality considering library complexity, strand-specificity, evenness, and continuity of annotated transcript coverage. We also evaluated the performance of the dU-Tn5 method in identifying nitrogen-responsive protein-coding genes and long non-coding RNAs in soybean roots. The results indicated that similar to 62-70% of differentially expressed genes detected from conventional libraries were also detected in dU-Tn5 libraries, indicating good agreement of our method with the current standard; moreover, their fold-changes were highly correlated (R>0.9). Thus, our method provides a promising 'do-it-yourself' stranded RNA-seq procedure for gene expression profiling. [GRAPHICS] .

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