盐藻3-磷酸甘油脱氢酶基因克隆

A STUDY ON GLYCEROL 3 PHOSPHATE DEHYDROGENASE GENE OF Dunaliella salina

采用Ｌａｍｂｄａ置换型载体ＥＭＢＬ３构建了盐藻基因组文库，文库大小为１．８×１０４，插入片段大小为１０～２０ｋｂ，从盐藻基因组文库中获得３个３－磷酸甘油脱氢酶基因的阳性克隆。

The genomic library(18,500 clones) of Dunaliella salina was obtained using a lambda replancement vector EMBL3 The main size of insertion is 10～20kb Comparing the conservative sequence encoding NAD binding region of glyoerol 3 phosphate dehydrogenase among Drosophila , rabbit and mouse, authors designed and synthesed a painr of primers The size of the primers was 21bp and 18bp, respectively PCR amplification was performed and the result of PCR showed a specific amplified region around 290bp(which is the same as in Drosophila DNA ) The recovered fragment was used as the probe for screening glyoerol 3 phosphate dehydrogenage gene from genomic library Nick translation system was used for labelling probe As a result of in situ hybridization, 3 positive clones were obtained Dot blot hybridization confirmed the result of in situ hybridization