本文選題：近交系小鼠 + 遺傳檢測��； 參考：《中國藥品生物制品檢定所》2005年碩士論文

【摘要】：H-2基因復合體是決定小鼠免疫遺傳品質最主要的基因群，其中以K、D基因區(qū)段產物的免疫反應性最強，最具決定性。因此建立小鼠H-2檢測技術，可保障和控制實驗小鼠的免疫遺傳質量，完善我國實驗動物遺傳檢測技術。 本研究分三個部分。首先建立了微量細胞毒法，引進小鼠主要組織相容性復合體國際標準單克隆抗體系列—H-2D~b(27-11-13)、H-2D~d(34-5-8)、H-2D~k(15-5-5.3)、H-2K~k(16-3.22.4)，和國產單克隆抗體H-2D~b(2)、H-2D~d(4)、H-2D~k(32)、H-2K~b(33)、H-2K~d(31)、H-2K~k(23)。制取小鼠脾細胞，加入特異的單克隆抗體，結果顯示抗原與特異抗體發(fā)生強烈的免疫反應，導致同型的脾細胞死亡，通過倒置顯微鏡觀察死亡細胞占總細胞的百分比來判定不同品系小鼠H-2基因型。同時進行了國內外單克隆抗體的對比研究，結果表明兩種抗體沒有顯著性差別。 實驗的第二部分，根據(jù)H-2D區(qū)和H-2K區(qū)的DNA序列，利用Megalin程序進行多序列比較，在各品系小鼠之間相同的區(qū)域內設計引物，擴增各品系小鼠之間不同的序列。H-2D區(qū)擴增片段為828bp，H-2K區(qū)擴增片段為104bp。經(jīng)過純化、連接、轉化、測序驗證擴增結果的準確性后，通過PCR方法擴增出特異區(qū)，從聚丙烯酰胺凝膠電泳圖譜中可觀察到不同品系的小鼠H-2基因型表現(xiàn)出不同的電泳條帶，從而可以區(qū)分不同的品系。 實驗的第三部分，為了提高實驗的靈敏度和特異度，，建立了微孔
[Abstract]:H-2 gene complex is the most important gene group which determines the immune genetic quality of mice. Therefore, the establishment of mouse H-2 detection technology can guarantee and control the immunogenetic quality of experimental mice and improve the genetic detection technology of laboratory animals in China. This study is divided into three parts. In this paper, the method of microcytotoxicity was first established, and the international standard monoclonal antibody series of major histocompatibility complex of mice, -H-2DBU 27-11-13C, H-2Ddc34-5-5-8, H-2DOKCU, 15-5-5-5.3, H-2DGK, 16-3.22.4H, and the domestic monoclonal antibody, H-2Db2Dd4, H-2Ddt4, were introduced. Mouse spleen cells were prepared and specific monoclonal antibodies were added. The results showed that the antigen reacted strongly with the specific antibodies, resulting in the death of the same type of spleen cells. The percentage of dead cells in total cells was observed by inverted microscope to determine the H-2 genotypes of different strains of mice. The results showed that there was no significant difference between the two antibodies. In the second part of the experiment, according to the DNA sequence of H-2D region and H-2K region, the multiple sequences were compared by Megalin program, and primers were designed in the same region among all strains of mice. The sequence of different strains of mice was amplified from 828bpnH-2K to 104bp2.The results showed that the amplified fragment of H-2D region was 828bp2D and the amplified fragment was 104bp2K. After purification, ligation, transformation and sequencing to verify the accuracy of the amplification results, the specific regions were amplified by PCR method. Different bands of H-2 genotypes of different strains of mice were observed from polyacrylamide gel electrophoresis atlas. Thus, different strains can be distinguished. In the third part of the experiment, in order to improve the sensitivity and specificity of the experiment, the micropore was established.
【學位授予單位】：中國藥品生物制品檢定所
【學位級別】：碩士
【學位授予年份】：2005
【分類號】：R-332

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