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中国农学通报

中国农学通报 ›› 2013, Vol. 29 ›› Issue (15): 157-163.doi: 10.11924/j.issn.1000-6850.2012-3573

所属专题： 生物技术

• 生物技术科学 • 上一篇    下一篇

盐生杜氏藻MAPK基因的克隆及表达分析

张婷 柴晓杰 王媛 姜玉声 丛玉婷   

  • 收稿日期:2012-11-02 修回日期:2012-12-22 出版日期:2013-05-25 发布日期:2013-05-25
  • 基金资助:
    国家自然科学基金;辽宁省教育厅科技研究项目

Cloning and Expression analysis of DsMAPK from Dunaliella salina

  • Received:2012-11-02 Revised:2012-12-22 Online:2013-05-25 Published:2013-05-25

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摘要： 采用RT-PCR和RACE技术克隆了盐生杜氏藻（简称盐藻）的一种促有丝分裂活化蛋白激酶基因（GenBank Accession JQ782412），命名为DsMAPK。对其进行生物信息学分析结果表明：该基因的cDNA全长为1861bp，开放阅读框（ORF）为1407bp，编码468个氨基酸，5′非编码区67bp，3′非编码区387bp；该蛋白为亲水性蛋白，无信号肽，无跨膜区域，为非跨膜蛋白，且定位于细胞质基质。二级结构预测表明该蛋白有25.2%的α-螺旋，12.4%的伸展片段，62.4%的自由卷曲；蛋白质同源性分析表明，与衣藻、团藻的亲缘关系最近。荧光定量PCR结果显示，在高盐胁迫（含3mol/L NaCl）下，DsMAPK基因的表达量显著上升，胁迫1h后表达量达到最大值，为正常水平（含1.5mol/ L NaCl）的5倍，差异达到极显著水平（P<0.01）。这些研究结果为进一步阐明DsMAPK的功能及作用机制奠定了基础。

关键词: 抗氧化, 抗氧化

Abstract: Dunaliella salina Mitogen-activated protein kinase gene (GenBank Accession No. JQ782412)（named DsMAPK）was cloned by RT-PCR and RACE technology for further bioinformatics analysis. The result shows that the full length of cDNA for DsMAPK is 1861bp, which contains 67 bp 5′UTR, 387bp 3′UTR and 1407bp open reading frame, coding 468 amino acids. The protein is a hydrophilic protein without signal peptide. According to our analysis, the protein belongs to a non-transmembrane protein category and is located in cytoplasm. Secondary structure prediction indicates that the protein consists of 25.2% a-helix, 12.4% strands and 62.4% random coils. Protein Drthology shows that the protein has a closest genetic relationship with those of Chlamydomonas reinhardtii and Volvox carteri f. nagariensis. By using real-time quantitative PCR ,we monitored the expression value of DsMAPK .The results show that the expression level of DsMAPK mRNA has significantly risen under high salt stress(including 3 mol/L NaCl), and it achieved the highest level after 1h dispose of 3 mol/L NaCl, which is 5 times larger than that of under 1.5 mol/L NaCl untreated condition(P<0.01). The research laid a foundation for further study of the function and working mechanism of DsMAPK.