Department of Soil and Crop Sciences, Texas A&M University, College Station, TX 77843-2474, United States.
Department of Soil and Crop Sciences, Texas A&M University, College Station, TX 77843-2474, United States.
Genomics. 2019 Dec;111(6):1517-1528. doi: 10.1016/j.ygeno.2018.10.009. Epub 2018 Oct 23.
Gene expression has been widely used in functional genomics research; however, the gene expressions quantified with different methods have been frequently inconsistent, thus challenging the conclusions from such research. Here we have addressed this issue, while taking into account RNA alternative splicing. We found that when a gene was subjected to RNA alternative splicing, it was impossible or difficult to properly quantify the expression of a transcript of the gene or its overall expression using quantitative real-time PCR (qPCR), Northern hybridization, microarray, or serial analysis of gene expression. Shot-gun RNA-seq was the most proper to quantify the expression of a transcript or a gene in such cases. Moreover, the expressions of individual transcripts quantified by shot-gun RNA-seq were highly reproducible (r = 0.90-0.98) between individuals. Therefore, shot-gun or full-length RNA-seq should be the method of choice to properly quantify the expression of a transcript or a gene.
基因表达已被广泛应用于功能基因组学研究；然而，不同方法定量的基因表达常常不一致，从而对该研究的结论提出挑战。在这里，我们解决了这个问题，同时考虑到了 RNA 可变剪接。我们发现，当一个基因发生 RNA 可变剪接时，使用定量实时 PCR(qPCR)、Northern 杂交、微阵列或基因表达系列分析技术，不可能或难以正确定量该基因的转录本或其整体表达。在这种情况下， shotgun RNA-seq 是定量转录本或基因表达的最合适方法。此外，shotgun RNA-seq 定量的单个转录本的表达在个体之间具有高度的可重复性(r=0.90-0.98)。因此，shotgun 或全长 RNA-seq 应该是正确定量转录本或基因表达的首选方法。
