MULT1转基因小鼠的基因型鉴定和表型的初步分析

The genotypic identification and elementary phenotypic analysis of MULT1-transgenic mice

目的对高亲和力NKG2D配体MULT1转基因小鼠进行基因型的鉴定和表型分析,并进行MULT1的初步功能研究。方法提取转基因小鼠的基因组DNA,PCR以鉴定转基因小鼠的基因型。提取小鼠尾部组织mRNA进行实时定量PCR,检测其基因转录情况。应用流式细胞分析仪及免疫组化对MULT1转基因小鼠多种组织细胞进行初步分析。结果获得鉴定阳性的MULT1转基因小鼠3 S品系25只,4 S品系13只。3 S和4 S品系的MULT1转基因小鼠的基因转录产物阳性,可用于进行下一步研究。MULT1主要表达在胸腺和小肠上皮间淋巴细胞(iIELs),且在胸腺和iIELs表面的表达与小鼠的年龄有关,而在脾脏细胞上基本不表达。iIELs流式细胞仪分析结果显示,CD8+T细胞比例降低,而CD4+CD25+调节性T细胞比例升高。结论经过基因型和表型的鉴定,表明已经成功构建MULT1转基因小鼠。对转基因小鼠表型的初步分析表明,MULT1可能与胸腺的发育、老化以及免疫调节相关。

Objective To identify high affinity ligand MULT1-transgenic mice,analyze its elementary phenotype,and to perform primary functional study in MULT1.Methods We identified the genotype of transgenic mice by PCR and detected transcription level of MULT1 in tails by real-time PCR.The flowcytometry and immunohistochemistry were used to analyze the expression of MULT1 in various tissues.Results 25 3＇S-strain and 13 4＇S-strain MULT1-transgenic mice were obtained and which contain high level of MULT1 transcripts.MULT1 protein is mainly expressed in intestinal intraepithelial lymphocytes（iIELs） and thymocytes but not on splenocytes.Moreover,the levels of MULT1 expression in iIELs and thymocytes are associated with age.Within iIELs,the percentage of CD8＋T cells decreased,while CD4＋CD25＋Treg cells increased,when compared with wild-type mouse.Conclusion We successfully produced MULT1-transgenic mice.MULT1 is possibly associated with thymic development and aging,as well as immunoregulation in mice.