大鼠脑谷氨酸脱羧酶基因的cDNA克隆及序列分析

胰岛β－细胞自身抗原蛋白之一是脑中谷氨酸脱羧酶（Ｇｌｕｔａｍｉｃａｃｉｄｄｅｃａｒｂｏｘｙｌａｓｅ，ＧＡＤ，ＥＣ４．１．１．１５）同源物。以双链ｃＤＮＡ为模板，用ＰＣＲ方法快速克隆了Ｗｉｓｔａｒ大鼠脑ＧＡＤ基因的ｃＤＮＡ，将此包括编码５９３个氨基酸的全长ＤＮＡ片段重组入ｐＵＣ质粒并用双脱氧末端终止法测定了全部序列，证明其全长为１７７９ｂｐ．经比较发现Ｗｉｓｔａｒ大鼠脑与Ｒｕｓｓｅｌｌ报导的大鼠脑ＧＡＤ基因序列，有一处碱基的差别，但并不涉及氨基酸的改变。同时还对用ＰＣＲ扩增长片段ＤＮＡ进行了方法学上的探讨。

The cDNA Cloning and Sequencing of Rat Brain Glutamic Acid Decarboxylase

By using double-strand cDNA (ds cDNA) synthesized from a purified poly(A)￣+-RNA population of rat brain,the cDNA encoding rat brain glutamic acid decarboxylase (GAD,EC 4. 1. l. 15), one of the autoantigens in β-cell, was amplified by PCR. The full-length cDNA fragment containing 1779 bp coding for 593 amino acids was inserted into PUC 12 vector and sequenced by the dideoxy chain termination method. It shows that only one base is different from those reported by Russell,which does not involve in change of amino acid. The method for amplifying long DNA fragment has also been developed.