非综合征唇腭裂（NSCL/P）是人类最常见的先天性畸形之一，其病因复杂。本课题组基于前期GWAS和大家系样本全外显子组测序研究，结合国内外研究进展，拟选择Arhgap29作为目标基因，利用CRISPR/Cas9基因编辑技术，建立Arhgap29条件性基因敲除小鼠模型。观察Arhgap29基因敲除后大体表型和组织学表现差异；应用转录组测序和蛋白质组学技术，筛选E9.5-14.5时期Arhgap29基因敲除后颌面部发育中差异表达的基因及蛋白，并通过原位杂交、RT-PCR、Western Blot检测进行验证；对获取的差异转录因子，应用CHIP-seq技术，定位转录因子所调控的靶基因，预测可能的信号通路并进行验证；同时，验证前期研究报道的IRF6-ARHGAP29-RhoA信号通路是否在Arhgap29基因敲除后小鼠异常表型中起作用。探索Arhgap29基因在颌面部发育及唇腭裂发生中的作用机制。

Non-syndromic cleft lip or palate (NSCL/P) is one of the most common congenital malformations of human beings, with complex etiology. Our previous researches have accomplished GWAS for NSCL/P of Chinese population, then launched whole exome sequencing for pedigree amples to obtain candidate genes. Combining with the domestic and foreign progress of the etiology of NSCL/P, Arhgap29 was proved the most likely pathogenic gene for NSCL/P. In order to illustrate the function of the gene and the possible pathogenic mechanism, this study intends to use the CRISPR/Cas9 mediated gene editing technology to establish Arhgap29 conditional knockout mice model, to observe the gross and histological phenotype in Arhgap29 gene knockout mice. Transcriptomic sequencing and proteomic studies were used to screen differentially expressed genes and proteins in the maxillofacial region after Arhgap29 knockout in the E9.5-14.5 mice. They were verified and screened by in situ hybridization, RT-PCR, Western blot detection. CHIP-seq was used to locate the target genes which were binding by the transcription factors in transcriptomic sequencing and proteomic studies. We will validate the reported IRF6-ARHGAP29-RhoA signaling pathway in Irf6 knockout mice. These will provide more evidence to explore the role of Arhgap29 in the mechanisms of development of maxillofacial region and its contribution to cleft lip or palate.