Gene expression in cornea of Balb/c and C57BL/6N mice

Gene expression was quanitified in 4 naive corneas from Balb/c and 4 corneas from C57BL/6N mice without intervention by RNAseq of total RNA with the Ovation Kit for model organisms. To avoid false positive differential expression from better alignment of the reads from C57/Bl6 mice to the reference representing a closely related strain while retaining the applicability of the standard reference genome annotation, two pseudogenomes were generated incorporating the known variants into the refernce and aligning to the resulting genomes. BAM files were then converted with Lapels to the standard reference, which includes conversion of genome coordinates and adjusting CIGAR strings. Then expression quantification is possible with respect to the standard gene model (here Ensembl version 94) again. Gene expression by sequencing of total RNA from cornea in two classical inbred mouse strains.本研究采用针对模式生物的Ovation试剂盒，通过总RNA的RNA测序（RNAseq）技术，对4只未受任何干预的BALB/c小鼠原生角膜以及4只未经过实验处理的C57BL/6N小鼠角膜的基因表达水平进行定量分析。为规避因C57/Bl6小鼠的测序读段（reads）与亲缘关系相近的参考基因组比对效率更高而产生的假阳性差异表达，同时保留标准参考基因组注释的适用性，研究人员生成了两个假基因组（pseudogenome）：将已知变异序列引入参考序列，并将测序读段比对至由此构建的基因组。随后，研究人员使用Lapels工具将BAM格式文件（BAM files）转换为适配标准参考基因组的格式，该转换过程包含基因组坐标的转换以及CIGAR字符串（CIGAR strings）的调整。至此，即可再次基于标准基因模型（本研究中为Ensembl版本94）完成基因表达定量分析。本数据集聚焦两种经典近交系小鼠的角膜组织，通过总RNA测序分析其基因表达情况。