hβ2m/HLA-B2704双转基因小鼠的制备及β2m对HLA-B2704表达的影响

制备hβ2m HLA B2 70 4双转基因小鼠 ,研究hβ2m对HLA B2 70 4表达的影响 .通过hβ2m转基因小鼠和HLA B2 70 4转基因小鼠交配 ,出生动物及后代经PCR初步筛选 ,采用Southern杂交对经PCR初步筛选的hβ2m HLA B2 70 4双转基因阳性小鼠基因组DNA标本作进一步鉴定 .阳性者进行RT PCR和流式细胞光度术检测其在mRNA和蛋白水平的表达 .hβ2m阳性小鼠和HLA B2 70 4阳性小鼠交配 ,生仔鼠 6 7只 ,其中 2 8只仔鼠同时整合hβ2m和HLA B2 70 4基因 .Southern杂交证实 ,阳性鼠同时都含有hβ2m和HLA B2 70 4基因 .阳性小鼠的皮肤、结肠、睾丸和脾脏组织中均有HLA B2 70 4和hβ2m基因的mRNA表达 ,其外周血淋巴细胞膜上HLA B2 70 4基因的蛋白表达为 35 87% ,在HLA B2 70 4单转基因小鼠外周血淋巴细胞膜上HLA B2 70 4基因的蛋白表达为 7 87% (Histogramsta tistics) .成功地制备了hβ2m HLA B2 70 4双转基因小鼠 .在hβ2m HLA B2 70 4双转基因小鼠的外周血淋巴细胞膜上 ,HLA B2 70 4基因在蛋白水平表达较高 ,而在HLA B2 70 4单转基因小鼠中则表达不明显 .hβ2m可与HLA B2 70 4重链分子结合 ,稳定和增高其在淋巴细胞膜上的表达

Production of hβ2m/HLA-B2704 Double Transgenic Mice and Effect of Human β2Microglobulin on Expression of HLA-B2704

To produce hβ2m/HLA\|B2704 double transgenic mice and to study the effect of human β\-2 microglobulin on the expression of HLA\|B27 molecule,human β\-2 microglobulin transgenic mice were mated with HLA\|B2704 transgenic mice.The presence of hβ2m/HLA\|B2704 gene in the founder mice and their progeny was confirmed by PCR.Further identification of hβ2m/HLA\|B2704 gene was determined by Southern blot hybridization.RT\|PCR and flow cytometry were carried out to detect the expression of hβ2m/HLA\|B2704 gene at mRNA and protein level.Kunming mice were used for produce transgenic mice.28 of 67(41\^8%) hybrid mice from hβ2m transgenic mice×HLA\|B2704 transgenic mice carried simultaneously human β\-2 microglobulin gene and HLA\|B2704 gene,which were confirmed by Southern blot hybridization analysis.Human β\-2 microglobulin gene and HLA\|B2704 gene were expressed in the skin,colon,testicle and spleen tissue of double transgenic mice at mRNA level.HLA\|B2704 gene was expressed on the cell membrane of their peripheral blood lymphocyte at protein level.The expression of HLA\|B2704 gene was 35\^87% in hβ2m/HLA\|B2704 double transgenic mice at protein level,but was 7\^87% in HLA\|B2704 transgenic mice (Histogram statistics).hβ2m/HLA\|B2704 double transgenic mice were produced successfully.The results suggest that hβ2m can combine with HLA\|B2704 heavy chain molecule to stabilize and increase the expression of HLA\|B2704 molecule on lymphocyte membrane at protein level.