Podnar Jessica, Deiderick Heather, Huerta Gabriella, Hunicke-Smith Scott
Genomic Sequencing and Analysis Facility, University of Texas at Austin, Austin, Texas.
Curr Protoc Mol Biol. 2014 Apr 14;106:4.21.1-4.21.19. doi: 10.1002/0471142727.mb0421s106.
This unit presents protocols for construction of next-generation sequencing (NGS) directional RNA sequencing libraries for the Illumina HiSeq and MiSeq from a wide variety of input RNA sources. The protocols are based on the New England Biolabs (NEB) small RNA library preparation set for Illumina, although similar kits exist from different vendors. The protocol preserves the orientation of the original RNA in the final sequencing library, enabling strand-specific analysis of the resulting data. These libraries have been used for differential gene expression analysis and small RNA discovery and are currently being tested for de novo transcriptome assembly. The protocol is robust and applicable to a broad range of RNA input types and RNA quality, making it ideal for high-throughput laboratories.
本单元介绍了用于从多种输入RNA来源构建针对Illumina HiSeq和MiSeq的下一代测序（NGS）定向RNA测序文库的方案。这些方案基于新英格兰生物实验室（NEB）用于Illumina的小RNA文库制备套装，尽管不同供应商也有类似的试剂盒。该方案在最终测序文库中保留了原始RNA的方向，从而能够对所得数据进行链特异性分析。这些文库已用于差异基因表达分析和小RNA发现，目前正在进行从头转录组组装测试。该方案稳健且适用于广泛的RNA输入类型和RNA质量，非常适合高通量实验室。
