Gene expression changes in the supraorbital cranial mesenchyme after beta-catenin deletion in vivo

Goal of this study was to examine gene expression changes upon conditional deletion of beta-catenin at e13.5 cranial mesenchyme. En1Cre;b-cateninfl/fl and En1Cre;b-cateninfl/+ supraorbital cranial mesenchyme was manually dissected from E13.5 embryos (4 mutants and 3 controls). RNA was isolated using Trizol and a Qiagen RNeasy Mini Kit. Libraries were prepared in the CWRU Genomics sequencing core using the Illumina TruSeq Stranded Total RNA kit-with Ribo Zero Gold. Paired-end sequencing was performed on an Illumina HiSeq 2500 v2 Rapid Run flowcell. The resulting 100bp reads were aligned to the mouse genome release mm9 genome using TopHat. Genomic assembly was completed using Cufflinks v1.3 (Roberts et al., 2011a, Roberts et al., 2011b, Trapnell et al., 2010, Trapnell et al., 2013). mm9_reFlat was used to annotate the data with a max intron length of 20,000 and genomic bias correction. Differential gene expression was determined with CuffDiff using the default settings plus genomic bias correction. contributor: Case Western Reserve University本研究旨在探究在胚胎日13.5（E13.5）颅间充质中条件性敲除β-连环蛋白（beta-catenin）后的基因表达变化。研究人员从13.5天龄小鼠胚胎中手动分离得到4例突变体（En1Cre;b-cateninfl/fl）与3例对照（En1Cre;b-cateninfl/+）的眶上颅间充质组织。总RNA采用Trizol试剂与Qiagen RNeasy Mini试剂盒进行提取。文库构建于凯斯西储大学（Case Western Reserve University）基因组测序核心实验室完成，使用Illumina TruSeq Stranded Total RNA试剂盒（搭配Ribo Zero Gold磁珠）完成建库。随后使用Illumina HiSeq 2500 v2 Rapid Run流动槽开展双端测序，获得100bp长度的测序读段。将所得读段通过TopHat软件比对至小鼠mm9版本参考基因组。基因组组装采用Cufflinks v1.3完成（引用Roberts等, 2011a、Roberts等, 2011b、Trapnell等, 2010、Trapnell等, 2013）。使用mm9_reFlat对测序数据进行基因注释，设置最大内含子长度为20000，并进行基因组偏倚校正。差异基因表达分析通过CuffDiff软件完成，采用默认参数并附加基因组偏倚校正步骤。本数据集的贡献方为凯斯西储大学（Case Western Reserve University）。