GGTA1基因敲除巴马小型猪的繁育及健康水平检测  

The Breeding and Health Level of GGTA1 Gene Knockout Bama Minipig(Sus scrofa)

作　　者：高景波 唐雨婷 龙川[2] 蒋应弟 石宁宁 冯冲[2] 潘登科[2] 刘霞[1] 

机构地区：[1]甘肃农业大学生命科学技术学院,兰州730070 [2]中国农业科学院北京畜牧兽医研究所/农业部畜禽遗传资源与种质创新重点实验室,北京100193

出　　处：《农业生物技术学报》2017年第10期1706-1713,共8页Journal of Agricultural Biotechnology

基　　金：国家重点基础研究发展计划(973计划)(No.2015CB554103);农业部畜禽遗传资源与种质创新重点实验室开放课题(No.nzdsys2015-1)

摘　　要：用于异种器官移植供体研究的α-1,3半乳糖苷转移酶基因(α-1,3-galactosyltransferase,GGTA1)敲除猪(Sus scrofa)的繁育性能和健康状态是异种器官移植之路能否成功的前提。为了观察本课题组制备的GGTA1基因敲除(GGTA1 knockout,GTKO)巴马小型猪(Bama minipig,BMP)的遗传基因型、窝产仔数及其血常规和血生化指标变化,评估其繁殖和健康状况,本研究对其进行定向培育,并繁育到F3代,通过PCR产物测序鉴定仔猪GGTA1基因敲除类型,异硫氰酸荧光素标记的植物凝集素(fluorescein isothiocyanate-griffonia simplicifolia isolectin B4,FITC-GSIB4)与仔猪外周血单个核细胞(peripheral blood mononuclear cells,PBMC)孵育后流式细胞术检测α-1,3半乳糖苷(α-1,3-galactosidase,α-1,3-Gal)的表达,以窝产仔数测定繁殖力,以血常规和血生化检测反映健康状况。结果显示,GGTA1基因的遗传符合孟德尔分离定律;GGTA1敲除纯合子(GGTA^(1-/-))仔猪的流式检测荧光强度低;GGTA^(1-/-)巴马猪母猪头胎窝产仔数为7.33±1.70头,经产母猪窝产仔数为9.43±1.68头,与已有报道的普通巴马小型猪繁殖力无明显差异;血常规和血生化检测的各项指标与普通巴马猪基本无差异。本研究建立了GTKO巴马小型猪家系,GTKO巴马小型猪遗传稳定,繁殖力正常,生理健康,GGTA^(1-/-)巴马小型猪有效解决了超急性排斥反应。The reproductive performance and health status of alpha-1, 3 galactosyltransferase gene（GGTA1） knockout pigs（Sus scrofa） is the key to success of xenotransplantation. Here we examined the genotype, litter size, blood routine, blood biochemical indexs of our group＇s GGTA1 knockout（GTKO） Bama minipigs（BMP） to assess their reproductive capacity and health status. GTKO pigs were mated with GTKO or wild type BMPs. The GTKO BMP have been bred to F3 generation. Firstly, the typing of GGTA1 gene knockout was identified by sequencing PCR products, α-1,3-Gal which is the product of alpha-1,3 galactosyl transferase gene was detected by flow cytometry after mixing Fluorescein isothiocyanate-Griffonia simplicifolia Isolectin B4（FITC-GSIB4） and peripheral blood mononuclear cells（PBMC）. Then, reproductive performance was evaluated by litter size. Health status was assessed by blood routine and blood biochemical indexes. The results showed that the inheritance of GGTAl gene was followed the Mendelian law; Low fluorescence expression were detected in GGTA1 double knocked （GGTAl＋） genotype. The litter size of gilt was 7.33±1.70, and the litter size of sow was 9.43±1.68, which had no significant difference in the fecundity with wild BMP Meanwhile, compared with wild-type BMP, no obvious difference were found in blood routine and blood biochemical indexes. In short, our results demonstrated that GTKO BMPs＇ family was established, with stable hereditary, normal fertility and healthy physiological indicators. The GGTA1/ Bama minipigs which effectively solved the hyperacute rejection, it can be used as ideal donor for xenotransplantation research.

关 键 词：α-1 3半乳糖苷转移酶基因(GGTA1) 巴马小型猪(BMP) 繁殖力 生理健康 

分 类 号：S828.99[农业科学—畜牧学]