Next Generation Sequencing Facilitates Quantitative Analysis of Murine Salivary Gland Transcriptomes at 8 and 32 weeks

Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular functions. The goal of this study is to compare NGS-derived salivary gland transcriptome profilings (RNA-seq) to better understand the molecular changes in adult sublingual glands in the absence of the Nkx2.3 transcription factors. Methods: Female sublingual salivary gland mRNA profiles were generated by deep sequencing, in 4 replicates for Nkx2.3 knockout mice, using Illumina. The sequence reads that passed quality filters were analyzed at the gene level with STAR followed by DESeq2. Results: Using an optimized data analysis workflow, we mapped about 20 million sequence reads per sample to the mouse genome (build mm9) and identified 496 genes that were differentially expressed between the wildtype and Nkx2.3-knockout murine female sublingual salivary glands . RNA-seq data provided valuable insights into the nature of the functional differences resulting from the Nkx2.3 disruption Conclusions: Our study represents the first detailed analysis of sublingal salivary gland transcriptome profiling differences resulting from the Nkx2.3 disruption, with biologic replicates, generated by RNA-seq technology. Our results confirmed functions of many previously studied genes. We conclude that RNA-seq-based transcriptome characterization would offer a comprehensive and sensitive evaluation of the gene expression. Female sublingual salivary gland mRNA profiles of 8- and 32-week old mice were generated by deep sequencing, in 4 replicates using Illumina.研究目的：下一代测序（Next-generation sequencing, NGS）已彻底革新了细胞功能的系统性分析范式。本研究旨在对比基于NGS获取的唾液腺转录组谱（RNA-seq），以深入解析Nkx2.3转录因子缺失时成年舌下腺的分子变化。 研究方法：本研究采用Illumina平台完成深度测序，对Nkx2.3基因敲除雌性小鼠的舌下唾液腺进行mRNA表达谱分析，每组设置4个生物学重复。对通过质量过滤的测序reads，采用STAR软件进行基因水平比对分析，随后使用DESeq2进行差异表达分析。 研究结果：通过优化的数据分析流程，我们将每个样本约2000万条测序reads比对至小鼠基因组（构建版本mm9），并鉴定出野生型与Nkx2.3基因敲除雌性小鼠舌下唾液腺间共496个差异表达基因。RNA测序（RNA-seq）数据为解析Nkx2.3基因敲除引发的功能差异本质提供了宝贵见解。 研究结论：本研究是首次通过RNA测序（RNA-seq）技术完成的、带有生物学重复的Nkx2.3基因敲除雌性小鼠舌下唾液腺转录组谱差异的详细分析。本研究结果验证了诸多此前已被研究的基因的功能。我们认为，基于RNA-seq的转录组表征可实现对基因表达的全面且灵敏的评估。 本研究通过Illumina平台完成深度测序，获取了8周龄及32周龄雌性小鼠舌下唾液腺的mRNA表达谱，每组设置4个生物学重复。