谷子DnaJ蛋白基因的克隆

Cloning of DnaJ-like Protein Gene from Foxtail Millet

DnaJ蛋白是近年研究较多的同热激等胁迫反应相关的基因。本研究用0.8%NaCl胁迫下的谷子幼苗提取RNA,用反转录酶M-MLV扩增得到第一链cDNA,再以第一链cDNA为模板进行PCR扩增,克隆获得了完整的谷子DnaJ蛋白基因,该基因cDNA全长1 260 bp,编码419个氨基酸,具有DnaJ蛋白分子伴侣系统的3个保守结构域。将谷子DnaJ蛋白基因构建入表达载体pGFP,获得了谷子DnaJ基因的表达载体。该基因的克隆和表达载体构建,为谷子DnaJ蛋白基因的功能分析以及谷子耐热抗旱机理研究有重要意义。

The DnaJ-like proteins and related genes are frequentily studied which are related with heatshock and other stresses in plant recentily. Using RT-PCR we cloned a foxtail millet Dna-J like gene in this report. RNA isolation was carried out by the Trizol method with 0.8 % NaCl solution treated foxtail millet seedling; first chain cDNA was obtained with the reverse transcripte enzyme M-MLV and then used as templated to PCR cloning the cDNA sequence of the foxtail millet Dna-J like genes. A PCR fragment of predicted size of 1.2 kb was cloned and sequenced. The cloned fragment contains 1 260 bp sequence which encodes a protein of 419 amino acids. Both the DNA sequence and amino acid sequence show great similarity with their corresponding sequences from rice, maize and wheat. The predicted protein has the three conserved domains which characteristic the structure of Dna-J protein family members. The cloned foxtail millet Dna- J gene was constructed into an expression vector for furthwer functional and transformation study.