枣树2-半胱氨酸氧化还原酶基因Zj2-CP的生物信息学分析及表达载体的构建

Bioinformatics Analysis and Construction of the Plant Expression Vector of Zizyphus jujube 2-Cys Peroxiredoxin(Zj2-CP)

本研究使用生物信息学相关软件对本课题组前期获得的枣树2-半胱氨酸氧化还原酶基因Zj2-CP(Gen Bank登录号:AB812086)的c DNA序列进行了分析。结果表明:该序列是一个全长为834 bp的开放阅读框,可编码277个氨基酸,分子质量为30.65 k D,理论等电点8.95,无信号肽和跨膜区,不属于分泌蛋白,有亲水性;二级结构以无规则卷曲为主,三级结构成弯曲螺旋状。氨基酸序列相似性分析表明,Zj2-CP基因编码的氨基酸序列与豇豆、玉米氨基酸序列的同源性最高,达到90%。以Zj2-CP基因的c DNA序列为模板进行PCR扩增,用内切酶消化后与载体PEZR(K)-LNY连接并转入大肠杆菌感受态细胞DH5α中,经酶切及测序证明,植物表达载体构建成功。研究结果可为深入探讨枣树2-半胱氨酸氧化还原酶基因的功能及枣树抗逆机制提供基础。

In our previous study, we obtained a c DNA sequence of Zizyphus jujuba 2-Cys peroxiredoxin Zj2-CP(DDBJ/EMBL/Gen Bank: AB812086). In this study, we used bioinformatics analysis tools for its' sequence analysis.The results showed that the full-length of the c DNA sequence was 834 bp and contained an open reading frame of277 amino acids. The protein's molecular weight was 30.65 k D and the theoretical PI was 8.95, hydrophilic, had neither signal peptide nor transmembrane domain. Secondary structure of Zj2-CP was mainly composed of random coil and the tertiary structure showed a forniciform helix structure. Amino acids homology analysis indicated that the amino acid sequences of Zj2-CP shared the highest 90% identity with Vigna radiate and Zea mays. Using the c DNA sequences of Zj2-CP as the template for PCR amplification, the PCR products were digested by restriction enzymes and connected to recombinant with plant expression plasmid PEZR(K)-LNY, then transferred the plant expression plasmid into the competent cells of E. coli DH5α. Restriction enzyme analysis and DNA sequences analysis demonstrated that the Plant Expression Vector was constructed successfully. These results provided a basisfor the further research of Zj2-CP's function and jujube's resistance mechanisms.