肺脏塌陷是导致新生克隆牛死亡、克隆效率降低的重要原因之一，肺泡表面活性蛋白SPB和SPC是维持肺泡容积稳定的重要蛋白，但新生克隆牛塌陷肺脏中SPB和SPC异常表达的调控机制还未见报道。在病理检查，比较cDNA表达谱，验证SPB和SPC基因异常表达的基础上：本研究将采用亚硫酸盐测序、染色质免疫共沉淀（ChIP）等技术研究新生死亡克隆牛塌陷肺脏中SPB和SPC基因启动子DNA甲基化、组蛋白修饰以及基因启动子区转录因子TTF-1的结合丰度；利用生物信息学技术及细胞转染技术预测并验证可能影响SPB和SPC蛋白生成的microRNA；采用蛋白质磷酸化芯片技术，研究EGFR信号通路磷酸化修饰对SPB和SPC基因表达的影响。从多角度、多层次研究塌陷肺脏中SPB和SPC基因异常表达的调控机制。本项目的实施，将为新生克隆牛肺塌陷分子机制的阐明和克隆牛生产效率的提高奠定重要的理论基础。

Alveolar collapse is a key cause of death of newborn cloned cattle and low efficiency of cloning. Pulmonary surfactant associated proteins, SPB and SPC, have a crucial role in maintaining regular alveolar volume. However, the regulatory mechanisms of SPB and SPC abnormal expression in collapsed lung of neonatal dead cloned cattle are rarely studied. On the basis of our studies on pathologic diagnosis, cDNA array data of collapsed and normal lung and verified differential expression of SPB and SPC, here we will try to elucidate the promoter DNA methylation, histone modifications and TTF-1 binding differences of SPB and SPC genes between collapsed lung of neonatal dead cloned cattle and healthy lung of age-matched normal cattle by bisulfite-sequencing and CHIP, predict and validate microRNAs that may regulate expression of SPB and SPC, study the influence of protein phorsphorylation of EGFR pathway on expression of SPB and SPC by phorsphorylation chip. So, regulation of SPB and SPC genes expression in collapse lung will be analyzed at mutil angles and mutil levels. We will contribute significantly to the elucidation of the mechanism of alveolar collapse of neonatal cloned cattle and the improvement of the efficiency of animal cloning by implementation of this program.