一、人體T細胞受器V基因片段使用情形之研究 二、人和老鼠T細胞受器Jα基因座內，一固定保留基因片段上結合因數之生化性質研究

Abstract

T細胞受器是T細胞辨認主要組織相容性複合體和抗原的覆合體的位置，而其多樣性產生的區域則是位於和免疫球蛋白上CDR3區域相當的位置。為了進一步瞭解多樣性產生得機制和T細胞受器V基因片段的使用情形，我們利用聚合?連鎖反應對人體胸腺和脾臟細胞的cDNA作增幅反應，然後作定序分析。結果發現，鹼基刪減和加入二種多樣性的產生大部分發生在J基因片段的5'端，較少發生在V基因片段的3'端；而且其長度也非常不一。另外，我們還發現有Vδ-Jα-Cα特殊轉錄產物出現，但是要進一步的實驗去證實他們是否有蛋白質的表現。 本論文的另一部分是關於固定序列基因塊（CSB）上結合因數的研究。CSB是位於T細胞受器Jα基因片段基因座內Jα3和Jα4之間，一段長125個bps的DNA片段，而且在老鼠和人之間有95％的序列相似性。早期的研究已經證實它是一段具有調節功能的基因片段。在本論文中，作者發現在老鼠胸腺和一T細胞株的細胞核萃取液內，有具特異性的結合蛋白能夠結合在CSB的二個區域中。由競爭性電泳膠遲緩試驗結果，發現這些結合能被固定的八聚體形式序列ATGCAAAT所競爭掉；經由序列比較，發現CSB上也有類似序列出現。當進一步用西南方轉漬法去研究時，發現這些片段除了彼此間有分子量相似的結合因數外，還有一些是屬於胸腺或T細胞株所特有的。下一步，我們將進行這些分子的純化和鑑定。

Most of the diversity in T cell receptor subunits resides in the region that is the equivalent of the CDR3 of Immunoglobulins. In order to learn more about the relative contributions of various mechanisms that generate this diversity and the usage of the TCR V gene segments, we used the polymerase chain reaction (PCR) to study TCR transcripts derived from human thymus and spleen cell and analysed the sequences of these transcripts. Most of the diversity is located at the 5'end of Jα genes segment. Deletion of bases from the ends of V and J genes does not occur with equal frequency. A greater number of bases were deleted or added on average from the ends of J genes. Special transcripts Vδ-Jα-Cα were found in thymus and spleen, further expertments are needs to clarify the expressional control of these transcripts. Conserved sequence block (CSB) is a DNA fragment with 125 bps that is located between Jα3 and Jα4 of the TCR Jα gene segment locus, with 95% sequence similarity between human and mouse. Previous studies have demonstrated the regulatory function of CSB. In this thesis, I found the specific DNA binding factors in the nuclear extract of mouse thymus and T-cell line that can bind to two regions of CSB. From EMSA competition assay, these factors can be competited by octamer motif ATGCAAAT but not other competitors. From South-Western blot, these fragments shared part of these binding factors with similar molecular weight, but some are specific for thymus or T-cell line. Further experiments will focus on the purification and identification of these factors.