發(fā)布時間：2018-02-01 21:47

  本文關鍵詞： 五指山豬 SLA復合體 單核苷酸多態(tài)性 氨基酸變異　出處：《華中農(nóng)業(yè)大學》2005年碩士論文　論文類型：學位論文

【摘要】：異種器官移植一直被認為是解決人體移植器官供體匱乏問題的方法。隨著轉(zhuǎn)基因及基因雙敲除豬的相繼問世,對異種移植的發(fā)展產(chǎn)生了巨大的推動作用。MHC編碼移植抗原,控制移植排斥反應,參與免疫應答調(diào)控和免疫識別,是移植物在受體內(nèi)能否成活的關鍵。鑒于此,本實驗首次比較詳細的完成了五指山小型豬群體中幾個經(jīng)典的SLA基因的SNPs檢測,并對五指山小型豬—人異種移植免疫進行初步的研究,獲得的結(jié)果如下: (1)采用PCR-SSCP結(jié)合克隆測序技術,檢測五指山豬群體中SLA-DQA,DQB第二外顯子的多態(tài)性。結(jié)果發(fā)現(xiàn)SLA-DQA第二外顯子存在堿基的顛換(A/C),引起氨基酸的改變(K/Q);而SLA-DQB第二外顯子區(qū)域沒有發(fā)現(xiàn)多態(tài)。 (2)利用AS-PCR技術對SLA—DQA進行準確分型。設計等位基因序列特異性引物直接從DNA水平上對存在多態(tài)的SLA-DQA基因進行分型,并取得了滿意的效果,與測序結(jié)果100%相符。 (3)依據(jù)豬EST所構建的EST重疊群,設計的豬特異引物分離并鑒定五指山豬群體中SLA-1,SLA-2基因第二和第三外顯子的多態(tài)性。在五指山豬群體中SLA-1,SLA-2基因分別存在兩個等位基因,并且這兩個基因多肽結(jié)合區(qū)與人相應部分氨基酸序列間均存在50%～55%的同源性。 (4)對五指山豬群體中SLA Ⅰ類基因核苷酸序列系統(tǒng)數(shù)分析,發(fā)現(xiàn)與SLA-3基因相比,SLA-1、SLA-2基因間存在較高的同源性。 (5)將豬SLA Ⅰ類基因與人HLA Ⅰ類基因NK細胞殺傷抑制受體結(jié)合區(qū)相應氨基酸進行了比較分析,結(jié)果發(fā)現(xiàn)它們都存在一個以上氨基酸的差異,這使得它們與NK細胞殺傷抑制受體基本上很難結(jié)合。
[Abstract]:Xenotransplantation has been regarded as a solution to the problem of donor shortage in human organ transplantation. With the development of transgenic and double-knockout pigs. For the development of xenotransplantation, MHC coding for the development of transplantation antigen, control transplant rejection, participate in the regulation of immune response and immune recognition. It is the key to the survival of grafts in vivo. In view of this, the SNPs detection of several classical SLA genes in Wuzhishan mini-pig population was completed in detail for the first time. A preliminary study on the xenotransplantation immunity of Wuzhishan mini-pig and human was carried out, and the results were as follows: 1) SLA-DQA in Wuzhishan pig population was detected by PCR-SSCP combined with cloning and sequencing. The polymorphism of exon 2 of DQB. It was found that the base transversion of exon 2 of SLA-DQA resulted in the change of amino acid. No polymorphism was found in the second exon of SLA-DQB. 2). AS-PCR technique was used to accurately type SLA-DQA. Allelic sequence specific primers were designed to type the polymorphic SLA-DQA gene directly from the DNA level. Satisfactory results were obtained, which were consistent with the sequencing results of 100%. According to the EST overlap population constructed by pig EST, the designed porcine specific primers were used to isolate and identify SLA-1 in Wuzhishan pig population. The polymorphism of exon 2 and exon 3 of SLA-2 gene. There were two alleles in SLA-1 and SLA-2 gene in Wuzhishan pig population. There was a 50% homology between the two polypeptide binding regions and the corresponding amino acid sequences of human. (4) the nucleotide sequence of SLA class I gene in Wuzhishan pig population was analyzed and compared with that of SLA-3 gene. There is high homology among SLA-2 genes. (5) the amino acids in the cytotoxic receptor binding region of porcine SLA class 鈪，

本文編號：1482943