Next Generation Sequencing Analysis of Wild Type and Rfx2-/- Testicular Transcriptomes at post natal day 21 [RNA-Seq P21]

Purpose: This study was carried out to determine the consequences of the Rfx2-/- genotype on spermatogenesis in the mouse Methods: RNA was extracted from decapsulated testes of 21 day old mixed background mice of either genotype. Deep sequencing was used to determine quantitative expression of the genomes from independent replicates of each genotype Results: RNA-Seq analysis identified some 105 genes that are down regulated at least 2-fold in Rfx2-/- testes, with ~50 being reduced at least 10-fold Conclusion: Spermatogenesis undergoes complete arrest just prior to the end of the round spermatid period of sperm development in mutant mice. Sequencing results showed that approximately 105 genes were downregulated 2 fold or more in the testes of mutant mice. Comparison of similar studies of targeted mutations in genes for other transcription factor demonstrate that Rfx2 has a large and nearly unique set of genes that depend on it directly or indirectly. A large number of downregulated genes are identified with cilia function. Testicular mRNA profiles were determined by deep sequencing using testes from 5 independent wild type and 6 independent Rfx2-/- mice研究目的：本研究旨在明确Rfx2-/-基因型对小鼠精子发生的影响。实验方法：从两种基因型的21日龄混合遗传背景小鼠的去被膜睾丸中提取总RNA，通过深度测序（deep sequencing）定量检测各基因型独立重复样本的转录组表达水平。实验结果：RNA测序（RNA-Seq）分析鉴定出约105个基因在Rfx2-/-小鼠睾丸组织中至少下调2倍，其中约50个基因的下调幅度至少达10倍。研究结论：突变小鼠的精子发生过程在精子发育的圆形精子细胞阶段结束前完全停滞。测序结果显示，突变小鼠睾丸组织中约105个基因的表达水平下调2倍及以上。通过与其他转录因子基因靶向突变的同类研究对比可知，Rfx2拥有一套规模庞大且近乎独特的、直接或间接依赖其调控的基因集合。大量下调基因与纤毛功能密切相关。本研究采用5只独立野生型小鼠与6只独立Rfx2-/-小鼠的睾丸样本，通过深度测序确定了小鼠睾丸的mRNA表达谱。