hLF转基因羊繁殖、遗传与表达稳定性

【目的】分析转基因对人乳铁蛋白（human Lactoferrin，hLF）转基因羊繁殖性能的影响，及转基因在不同世代中的遗传与表达稳定性。【方法】应用自主研发的原代hLF转基因山羊（G0代公羊）建系，与普通奶山羊常规交配获得F1代；F1代公羊与F1、F2代母羊或普通奶山羊（N）交配获得F2代；F2代公羊与F2代母羊交配获得F3代。分别采用PCR和ELISA方法对其后代个体进行整合与表达鉴定；统计各世代不同配种方式所获得的后代数量、转基因整合率、母羊受胎率、羔羊异常率等繁殖性能数据。【结果】经过多个繁殖周期，各世代转基因羊受胎率、产羔率等繁殖性状均正常。获得F1代hLF羊19只，转基因整合率为24.7%（19/77）；F2代hLF羊86只，其中通过（F1♂×N♀）交配方式获得的是62只，转基因整合率为33.2%（62/187）；通过（F1♂×F1♀）交配方式获得hLF羊16只，转基因整合率为76.2%（16/21）；通过（F1♂×F2♀）交配方式获得hLF羊8只，转基因整合率为34.8%（8/23）；获得F3代hLF羊9只，转基因整合率为52.9%（9/17）。ELISA方法测定F1、F2、F3代羊泌乳20周乳汁中hLF的表达发现，hLF在转基因羊乳汁中稳定表达，3个世代hLF羊乳中的平均表达量分别为351.75、340.00和326.25 μg•mL-1。【结论】hLF基因的转入未对转基因羊群的繁殖性能产生显著影响，hLF基因能在山羊群体世代间稳定遗传和表达。

Breeding and Expression Stability of Human Lactoferrin Transgenic Goat

【Objective】This study was conducted to analyze the effects of transgenosis on the reproduction of human lactoferrin(hLF) transgenic goat,and the genetic and expression stability of exogenous gene in each generation of transgenic animals.【Method】Family line was established with self-developed hLF transgenic goats(G0♂),F1 generation was composed by integrated offspring individuals of G0♂ mated with ordinary dairy goat,F2 was generated by F1 males mated with F1,F2 or ordinary females,and F3 was generated by F2 males mated with F2 females.PCR and ELISA were implied to identify the integration and expression of exogenous gene in offsprings.Reproductive data such as quantity of offspring with different mating patterns,rate of transgenic integrated individuals,conception rates,abnormal rates of lamb in each generation were counted statistically.【Result】After several reproductive cycles,no abnormalities of conception rates and lambing rates were observed.The results showed that,there were 19 hLF transgenic individuals in F1 generation,with integration rate of 24.7%(19/77),86 individuals in F2 generation,in which 62 were generated by F1♂×N♀,16 were obtained by F1♂×F1♀,and 8 were obtained by F1♂×F2♀,with the rate of 33.2%(62/187),76.2%(16/21) and 34.8%(8/23),respectively,9 hLF transgenic individuals in F3 generation and the integration rate of F3 was 52.9%(9/17).ELISA was conducted to measure the expression of hLF in F1,F2 and F3 in 20 weeks,and the results showed that hLF expressed stably in each transgenic generation and the average expression level in milk were 351.75,340.00 and 326.25 μg·mL-1,respectively.【Conclusion】The results suggested that,there is no significant effect of transgenosis on reproductive performance of hLF transgenic goat,and exogenous hLF gene could inherit and stably express in each generation of goats.