论文部分内容阅读
有研究显示,组蛋白去乙酰基酶复合物(BRAF-HDAC complex,BHC)组分BHC80基因敲除导致小鼠出生一天内死亡。为此,本研究在斑马鱼中下调BHC80基因表达,以深入研究其在生物体发育中的作用。根据斑马鱼BHC80芋列,设计合成吗啡啉修饰的反义寡核苷酸,并将其显微注射到单细胞或双细胞期的野生型胚胎中,并用RT-PCR方法验证其有效性。分析BHC80基因阻抑后对胚胎发育,尤其心脏表型和功能的影响。结果显示,吗啡啉修饰的反义寡核苷酸有效下调BHC80基因表达,其对胚胎发育异常的影响呈剂量依赖性。BHC80基因表达下调的斑马鱼胚胎心脏出现多种异常的表型,包括心房心室大小异常、环化不完全、严重者心脏发育呈管状、心搏减弱,心率减慢、心室收缩分数降低。结果表明,BHC80基因表达下调致使胚胎心脏发育异常,心功能受损,这可能是BHC80基因敲除哺乳动物出生后很快死亡的重要原因之一,为进一步阐明心脏发育机制提供了很好的研究工具。
Some studies have shown that BHC80 gene knockdown of the BRAF-HDAC complex (BHC) component causes the mice to die within one day of birth. For this reason, this study down-regulated BHC80 gene expression in zebrafish in order to further study its role in the development of the organism. According to the zebrafish BHC80 gene array, a synthetic morpholine modified antisense oligonucleotide was designed and microinjected into the single-cell or two-cell wild-type embryos, and its effectiveness was verified by RT-PCR. The effect of BHC80 gene repression on embryonic development, especially cardiac phenotype and function, was analyzed. The results showed that morpholine modified antisense oligonucleotides effectively down-regulated BHC80 gene expression, and its effect on embryonic development in a dose-dependent manner. The aberrant phenotypes of the embryonic heart of zebrafish with down-regulated BHC80 gene expression include abnormal atrial ventricular size, incomplete circularization, severe cardiac development, decreased heartbeat, slowed heart rate and decreased ventricular systolic scores. The results showed that the downregulation of BHC80 gene resulted in abnormal embryonic cardiac development and impaired cardiac function, which may be one of the important causes of death of BHC80 knockout mammals after birth, which provides a good study for further elucidating the mechanism of cardiac development tool.