鸡与鹌鹑属间杂交早期胚胎bcl-2、P53基因表达的差异及发育性变化

The Expresstional Differences and Developmental Changes of bcl-2, P53 Gene in Early Embryos of Generic Hybrids of Chicken-Quail

【目的】探讨细胞凋亡因子bcl-2、P53对鸡与鹌鹑属间杂交早期胚胎发育的影响。【方法】人工授精获得鸡(♂)与鹌鹑(♀)杂交种蛋,按照鸡标准孵化条件同批入孵,随机采集66、72、78、84、90、96、102、108、114、120h活胚。采用RT-PCR方法,用Wpkci和β-actin引物进行多重PCR鉴定胚胎样本性别,后选取各时间点雌、雄胚胎各4枚,以β-actin为内标,测定bcl-2、P53的mRNA相对丰度。【结果】(1)雄性胚胎66～114hbcl-2mRNA表达维持在较低水平,96h有所降低,随后回升到原水平,120h极显著升高(P<0.01),达到峰值;雌性胚胎66～114hbcl-2mRNA表达维持在较低水平,114h极显著升高(P<0.01),此后维持在较高水平,120h达到峰值。不同日龄之间的比较,114h雌性表达显著高于雄性(P<0.05);(2)雌、雄胚胎P53mRNA发育性表达模式基本一致。66h表达水平较高,72h表达降低,达到低谷,雄性达到显著(P<0.05),雌性整体差异不显著,随后上升维持在原水平;不同日龄之间表达差异不显著。【结论】114h雌性胚胎bcl-2mRNA表达高峰点与其P53bcl-2表达最低水平相吻合;72h雌、雄胚胎P53mRNA表达与P53bcl-2表达水平一致,处于低谷点;72、114h早期杂交胚胎bcl-2、P53基因表达出现异常。

[Objective] To investigate the influences of apoptotic factors bcl-2, P53 on early embryos of generic hybrids of chicken-quail. [Method] The cross-bred eggs of quail （♀） acquired artificially inseminated by chicken（♂）-semen, hatched the eggs in one batch according to the standard condition of chicken, and collected the early living-embryos of 66, 72, 78, 84, 90, 96, 102, 108, 114 and 120 h at random. The method of RT- PCR was used to multiply PCR to identify and clarify the quail＇s sex and embryo sex with primers of Wpkci and β-actin, then selected four embryos of female and male respectively at each period, taken β-actin for internal standard to determine the relative quantity of embryo bcl-2 and P53 mRNA. [Result] （1） The bcl-2 mRNA expression of 66-114 h of male embryos maintained lower level, declined on 96 h, then reached the initial level, rose distinctly on 120 h （P〈0.01） and reached the peak; the bcl-2 mRNA expression of 66-114 h of female embryos maintained lower level, rose distinctly on 114 h （P〈0.01）, then maintained higher level and reached the peak on120 h; the female bcl-2 mRNA expression of 114 h was significantly higher than male of the same days （P〈0.05）; （2） The patterns of P53 mRNA expression of male and female embryos were basically consistent. They were higher on 66 h, declined on 72 h （P〈0.05） and reached the lowest level, declined distinctly in male, but the whole differences were not significant in female, then reached and maintained the initial level. Compared the expression between different sexes, the differences were not significant. [Conclusion] At 114 h the expressional peak of bcl-2 mRNA of female embryos coincide with its P53/bcl-2 lowest expressional level; At 72 h P53 mRNA expressional quantity of male and female embryos were consistent with their P53/bcl-2 expressional level, both were at low level. At 72, 114 h the bcl-2, P53 mRNA expression of early embryos of hybrids were abnormal.