IGF-I及其受体IGF-IR基因在鸡胚胎腿肌发育过程中表达模式分析

Expression of IGF-I and IGF-IR mRNA in leg muscle during chicken embryonic development

为研究胰岛素样生长因子IGF-I及其受体IGF-IR在鸡胚胎腿部肌肉发育过程中的作用,选取藏鸡和矮小蛋鸡受精蛋孵化后,采集鸡胚胎孵化第9天、第13天和第20天时腿部肌肉组织,并利用荧光定量PCR技术检测IGF-I和IGF-IR mRNA的表达水平,同时检测IGF-I基因编码区的单核苷酸多态性并利用RFLP(限制性片段长度多态性)方法对筛选到的单核苷酸多态性进行群体水平分型。结果表明:藏鸡和矮小蛋鸡胚胎腿部肌肉组织在孵化第9天和第13天时的IGF-I和IGF-IR的mRNA表达量均极显著高于第20天,且藏鸡IGF-I的mRNA表达量在第9天也显著高于第13天,而矮小蛋鸡的IGF-I mRNA表达水平在第9天和第13天无明显差异。此外,在整个孵化过程中,矮小蛋鸡的IGF-IR的表达模式与IGF-I表达模式基本一致。而藏鸡IGF-IR的表达水平在第9天明显高于第13天但未达到显著水平(P>0.05),同时发现第9天藏鸡IGF-IR的表达水平也高于矮小蛋鸡(P>0.05)。除此之外,在藏鸡和矮小蛋鸡的IGF-I的编码区均发现c.282A>G单核苷酸多态性,该SNP在藏鸡中主要以杂合子AG基因型为优势基因型,基因型频率为0.57,而在矮小蛋鸡中主要以GG基因型为优势基因型,基因型频率为0.77。根据试验结果分析得出,IGF-I和IGF-IR在腿部肌肉组织的表达水平在鸡胚胎孵化中期(第9天和第13天)显著高于孵化后期(第20天),可能是由于中期属于胚胎腿部肌肉快速发育阶段而后期腿部肌肉发育变得相对迟缓,同时IGF-I和IGF-IR的表达水平也随之发生变化。

To illustrate the function of IGF-I and IGF-IRin leg muscle during chicken embryo development,fertilized eggs from Tibet chickens and Dwarf chickens were hatched and the leg muscle tissues of chicken embryo were collected at day 9,13 and 20.Real-time PCR was used to detect IGF-I and IGF-IR mRNA expression level.Furthermore,coding region of IGF-I was sequenced for searching the single nucleotide polymorphism（SNP）and then identified SNP was genotyped with RFLP in chicken population.The results indicate that the mRNA level of IGF-I and IGF-IR was significant higher in both day 9and day 13 than day 20.The IGF-I mRNA expression was also significant higher at day 9in relative to day13 in Tibet chicken while no obvious difference of the IGF-I mRNA was observed between day 9and day 13 in Dwarf chicken.Furthermore,the expression pattern of IGF-I is similar to the IGF-IR duringthe Dwarf chicken incubation.The expression level of Tibet IGF-IRis higher in day 9than day 13,but no significant（P0.05）.At day 9,the mRNA level of Tibet IGF-IR is higher than Dwarf chicken（P0.05）.In additional,a single nucleotide polymorphism c.282AG was detected in coding region between Tibet and Dwarf chicken.In Tibet chicken,AG genotype was dominant which the frequency is 0.57 and GG genotype was dominant in Dwarf chicken which the frequency is 0.77.This study found that the mRNA level of IGF-I and IGF-IRin chicken leg muscle were significant higher at middle stage（day 9and day 13）than late stage（day 20）.The potential reason behind it is that development of leg muscle is faster at middle stage than late stage resulting to the mRNA level of IGF-I and IGF-IR changed.