中华鳖MHC I α2链基因克隆及序列分析

Molecular cloning and sequencing analysis of MHC class I alpha 2 domain from soft shelled turtle(Trinyx sinensis)

目的：为了探明中华鳖ＭＨＣ的结构与功能和寻找该种群的分子标记，对中华鳖ＭＨＣｃｌａｓＩα２基因中由两个半胱氨酸形成二硫键的区域进行了克隆和序列分析。方法：根据人和动物的ＭＨＣＩａα２链中两个半胱氨酸侧翼的保守序列设计了混合型引物，采用ＰＣＲ法从中华鳖基因组ＤＮＡ中克隆了ＭＨＣＩα２链（ＴｒｓｉＢＸ１）。结果：经氨基酸序列的同源性分析，ＴｒｓｉＢＸ１存在抗原多肽结合位点（Ｔ１４３、Ｋ１４６、Ｗ１４７、Ｙ１５９）以及β２微球蛋白结合位点（Ｑ１１５、Ａ１１７、Ｄ１１９、Ｇ１２０、Ｄ１２２），并与人和动物的ＭＨＣＩａα２存在４１１％～６３９％同源性。结论：ＴｒｓｉＢＸ１属ＭＨＣｃｌａｓｓＩａ类基因，可作为中华鳖种群的分子标记。

Objective:In order to research the MHC in soft shelled turtle and looking for a specific marker in vertebrate.Methods:The MHC class I α2 domain clone(Trsi BX1)was isolated from soft shelled turtle by PCR using mixed primers based on two highly conserved amino acid(aa)sequence blocks surrounding two cysteine residues in the second domain of MHC class Ia α2 chains of humans,mice,chickens and lizard.The aa sequence similarities between Salmo salar and Xenopus,and lizard,mouse and human.Results:The data indicate that four residues into the antigenic recognition site(T 143、K 146、W 147、Y 159)are conserved,and the β2 microglobulin binding sites (Q 115、A 117、D 119、G 120、D 122) are also conserved in the Trsi BX1.Conclusion:The Trsi BX1 is part of the MHC class Ia and may identified in vertebrate using specific marker.