目的 克隆红花花瓣中bZIP20(Basic region/leucine zipper motif)基因,研究其在不同组织中的表达量并构建其植物表达载体。方法 根据红花转录组测序结果挑选bZIP基因的设计引物,以红花花瓣总RNA为模板,采用RT-PCR法扩增bZIP20基因开放阅读框(ORF)片段,利用RT-PCR法分析在红花不同组织以及尖孢镰刀菌侵染后红花根部bZIP20基因的表达量,同时构建植物表达载体pBASTA-bZIP20。结果 bZIP20基因ORF长981 bp,编码326个氨基酸(GenBank登录号为KT692605)。红花bZIP20与其他物种氨基酸具有一定的同源性,其与芝麻、野茶树的氨基酸序列相似性高达85.41%和83.99%。实时荧光定量PCR分析表明,bZIP20基因在不同组织中的表达水平具有显著差异,在花中呈现高表达,而在其他组织中低表达。接种尖孢镰刀菌的红花根部组织中bZIP20基因的表达显著上调。结论 成功地对bZIP20基因进行克隆及表达分析,并构建植物表达载体pBASTA-bZIP20。

Objective To clone bZIP20 (basic region/leucine zipper motif) gene from Carthamus tinctorius, analyze the expression level in different plant tissues, and construct the plant expression vector. Methods The bZIP20 gene was cloned by RT-PCR techniques, and the protein characteristics were analyzed by bioinformatics, and phylogenetic tree was constructed. The expression of bZIP20 gene in different tissues and the roots after inoculated by Fusarium oxysporum were analyzed using real time-PCR, and the plant expression vector pBASTA-bZIP20 was constructed. Results The ORF sequence of bZIP20 gene was 981 bp, encoded a protein of 326 amino acids (GenBank: KT692605). Sequence alignment and phylogenetic tree analyses showed that bZIP20 had 85.41% and 83.99% of consistency with bZIP of Sesamum indicum and Camellia assamica. Real-time PCR results showed significant differences, the highest expression level of bZIP20 gene was detected in flower, and was highest in the bud period, bZIP20 gene was significantly increased in root tissue inoculated with F. oxysporum. The plant expression vector pBASTA-bZIP20 was obtained. Conclusion The bZIP20 gene of safflower is successfully cloned, and the expression is analyzed. The plant expression vector pBASTA-bZIP20 is constructed.

国家高技术研究发展计划(“863”)项目(2011AA100606);国家自然科学基金资助项目(31201237);吉林农业大学大学生创新创业项目(201410193045);吉林省教育厅“十三五”科学技术研究项目重点项目(吉教科合字[2016]179号);吉林省科技厅中青年领军人才及优秀创新团队项目(20111815)