Gene expression at E9.5 after deletion of type I BMP receptors in the mouse lens placode

The type I BMP receptors, Bmpr1a and Acvr1 were conditionally deleted from the mouse lens placode with Le-Cre. The heads of wild type and knockout E9.5 mouse embryos were laser micodissected to isolate the lens ectoderm from three embryos of each genotype. Total RNA was purified and reverse transcribed and amplified using a NuGEN Ovation Pico WTA system kit. cDNA was biotinylated and hybridized to Illumina Mouse6 V1.1 bead arrays. Three wild type and three knockout embryos were used. RNA obtained from the lens ectoderm was pooled, amplified and used for microarray analysis. Statistical analysis was performed using the values from the 25-50 beads on each array.本研究借助Le-Cre重组酶，从小鼠晶状体基板中条件性敲除I型骨形态发生蛋白受体（Type I BMP Receptors）Bmpr1a与Acvr1。选取胚胎发育第9.5天（E9.5）的野生型与基因敲除型小鼠头部各3枚，通过激光显微切割（laser microdissection）技术分离两种基因型胚胎的晶状体外胚层。纯化总RNA后，采用NuGEN Ovation Pico WTA系统试剂盒完成逆转录与扩增。将所得互补DNA（cDNA）进行生物素标记，随后与Illumina Mouse6 V1.1微珠芯片阵列进行杂交。将各样本获取的晶状体外胚层RNA混合后进行扩增，用于基因芯片分析。统计分析基于每张芯片上25至50个微珠的信号值开展。