腹水综合征肉鸡肺脏缺氧诱导因子-1α基因克隆及表达研究

Cloning and Expression of Hypoxia-inducible Factor 1α Gene from Ascitic Broilers' Lung Tissue

从21日龄患腹水综合征(AS)鸡肺脏中提取RNA,根据健康鸡胚心室肌细胞缺氧诱导因子-1α(HIF-1α)基因序列(登录号为NM204297)设计引物,采用RT-PCR技术进行cDNA扩增,扩增产物经琼脂糖凝胶电泳分离、纯化、回收后与载体pMD18-T连接,转化大肠杆菌DH5α,筛选阳性克隆,测序。结果表明,已成功克隆的阳性重组子经测序鉴定其片段大小与预期大小完全一致,为1 576 bp。序列同源性分析比较表明,该cDNA及其推导的氨基酸序列与鸡胚HIF-1αcDNA和氨基酸序列同源性分别为99%和98%。采用RT-PCR技术检测4只21日龄正常肉鸡和4只21日龄AS患鸡肺脏HIF-1αmRNA,结果表明,AS患鸡肺脏HIF-1αmRNA表达丰度极显著高于正常肉鸡(P<0.01)。本试验证明,HIF-1α与肉鸡肺动脉高压导致的腹水综合征相关。

The expression of hypoxia-inducible factor 1α gene（HIF-1α） in the lung of ascitic broilers was studied in comparison with healthy broilers, cDNA encoding HIF-1α from the lung of as citic broilers was transfected into E. coli DHSα by using pMD18-T vector. Sequence analysis showed that the cloned 1 576bp cDNA and the deduced amino acid sequence shared 99% and 98% homology respectively with chicken embryo HIF-1α cDNA. The expression of HIF-1α in the lung of 4 healthy broilers and 4 ascitic broilers were then examined by RT-PCR. Results showed HIF-1α mRNA abundance was significantly higher in the lung of ascitic broilers than that in healthy broilers（P〈0.01）. The result suggested that HIF-1α may be involved in the development of ascitic symptom induced by pulmonary hypertension of broilers.