猪4号染色体产肠毒素型大肠杆菌F41受体基因的鉴别研究

Enterotoxigenic Eschoerchia coli (ETEC) F41 is one of major pathogen causing diarrhea in neonatal piglets，resulting in significant economic loss in the pig industry.The pathogenicity of ETEC F41 depends on whether F41 fimbriae would adhere to their specific receptors on brush borders of enterocytes, and the key point to conduct breeding for disease resistance of ETEC F41 is to isolate and identify the gene encoding ETEC F41 receptor. In our previous study, the ETEC F41 receptor gene had been fine mapped to a region of 2.42 Mb between SW2509 and KVL2754 on pig chromosome 4 (SSC4). This project would develop high density SNP markers in the 2.42 Mb-region and genotype these markers on 400 samples from the Sutai bred with TaqMan OpenArray Genotyping assay, leading to a narrowed region of the gene encoding ETEC F41 receptor through linkage disequilibrium association analysis. Positional candidate genes will be identified according to the results from fine mapping, human-pig comparative map and the properties of ETEC F41 receptor. The candidate genes will be resequenced to identify SNPs, and further association analyses would be performed in a outbred population to identify the target gene and its causative mutation.These findings would provide novel technique to select for disease resistance to ETEC F41 in the pig industry.

产肠毒素大肠杆菌（ETEC）F41是引发新生仔猪腹泻病的主要病原菌之一，给养猪生产造成巨大经济损失。ETEC F41的致病性取决于其是否与猪小肠上皮细胞表面的特异性受体结合，因而，分离和鉴别猪ETEC F41受体基因是抗F41仔猪腹泻病育种的关键。项目申请人前期已将猪ETEC F41受体基因定位于4号染色体标记SW2509至KVL2754之间约2.42 Mb的区域。本项目拟在此区域开发高密度SNP，定制分型芯片，采用TaqMan OpenArray基因分型技术扫描400只苏太猪群体，通过基于连锁不平衡的关联分析进一步缩小ETEC F41受体基因定位区域。综合精细定位结果、比较基因图谱、受体的理化特性等筛选目的基因并对其进行重测序SNP搜寻，在远缘群体中开展关联分析以鉴别目的基因及其因果突变位点。本研究结果将为猪ETEC F41侵染腹泻的抗病育种提供理论依据和新型技术。

产肠毒素大肠杆菌（ETEC）F41引发的新生仔猪腹泻病是养猪生产中的常见病，给养猪业造成了巨大经济损失。本研究在课题组已将猪ETEC F41受体基因初步精细定位在4号约2.42 Mb的区域基础上，利用猪60K芯片扫描苏太猪群体及白色杜洛克×二花脸资源家系，采用全基因组关联分析及连锁不平衡的关联性分析，将ETEC F41受体基因定位区域进一步缩小至标记ALGA0022658（位于6.95 Mb处）和ASGA0017783（位于7.16 Mb处）间的1.21-Mb区域。根据人-猪比较基因组学及ETEC F41受体理化特性，选择该区域内ST3GAL1和ZFAT基因作为ETEC F41受体的重要位置候选基因开展研究。对这2个基因进行了全长cDNA的克隆、SNP的搜寻及鉴定、选择了部分SNP在资源群体和中、西方猪种远缘群体中使用SNaPSHOT技术进行基因分型，并与ETEC F41黏附表型进行关联性分析，鉴别到5个与ETEC F41易感性之间的关联性达到了显著水平的分子标记。本研究不仅证实了与ETEC F41侵染相关的主效QTL位于猪4号染色体上，而且进一步将目标区域缩小至标记ALGA0022658-ASGA0017783（6.95 Mb-7.16 Mb）间，为最终确定ETEC F41受体目的基因及其因果突变奠定了重要的前期工作基础。通过对位置候选基因ST3GAL1和ZFAT基因的克隆及多态性分析，鉴别到的与ETEC F41黏附表型显著相关的位点为利用标记辅助选择（MAS）开展抗ETEC F41腹泻病分子遗传育种提供了很好的分子标记。本研究成果，在国际期刊Animal（影响因子1.648）发表SCI文章一篇，申请专利一项（申请号201510324439.4）。