红花二氢吡啶二羧酸合酶基因片段的分离及表达分析

发布时间：2018-07-06 13:11

本文选题：红花 + 二氢吡啶二羧酸合酶　；参考：《中草药》2017年02期

【摘要】：目的克隆红花Carthamus tinctorius赖氨酸合成途径关键酶二氢吡啶二羧酸合酶(dihydrodipicolinate synthase,DHDPS)基因并研究其在不同发育时期红花籽粒中的表达量。方法根据红花转录组文库注释信息筛选出与红花DHDPS(Ct DHDPS)基因相关的Unigenes,设计引物,以红花总RNA的反转录产物为模板,采用RT-PCR技术克隆Ct DHDPS基因片段,连接p EASY-T1克隆载体,经PCR和酶切鉴定,筛选阳性克隆并测序。同时利用荧光定量PCR技术对其在红花籽粒不同发育时期基因表达量进行分析。结果分离到长度为396 bp的Ct DHDPS基因片段,系统发育树分析表明该基因与其他物种的DHDPS基因具有较高的同源性。结论克隆了Ct DHDPS基因的核心片段,根据Ct DHDPS基因片段设计引物,对不同品种不同发育时期红花种子进行基因表达量分析,Ct DHDPS基因在红花品种川红1号初花后14 d表达量最高。
[Abstract]:Objective to clone the dihydrodipicolinate synthase (dihydrodipicolinate synthase) gene from Carthamus tinctorius lysine biosynthesis pathway and to study its expression in Carthamus tinctorius grains at different developmental stages. Methods Unigeneses related to DHDPS (Ct DHDPS) gene were screened according to the annotated information of Carthamus transcriptome library. Primers were designed. The fragment of Ct DHDPS gene was cloned by RT-PCR and cloned into pEASY-T1 clone vector using reverse transcription product of total RNA of safflower as template. The positive clones were screened and sequenced by PCR and enzyme digestion. At the same time, fluorescence quantitative PCR technique was used to analyze the gene expression in different development stages of safflower grain. Results A 396bp fragment of Ct DHDPS gene was isolated. Phylogenetic tree analysis showed that this gene had high homology with DHDPS gene of other species. Conclusion the core fragment of Ct DHDPS gene was cloned and primers were designed according to the fragment of Ct DHDPS gene. The expression of Ct DHDPS gene in different varieties and different development stages of safflower seeds was analyzed. The expression level of Ct DHDPS gene was the highest 14 days after the early flowering of Ct DHDPS gene.
【作者单位】：吉林农业大学生物反应器与药物开发教育部工程研究中心;吉林农业大学中药材学院;吉林农业大学生命科学学院;
【基金】：国家高技术研究发展(“863”)计划(2011AA100606) 国家自然科学基金资助项目(31401445) 吉林省科技厅医药产业推进计划项目(20140311034YY) 教育部博士点基金(20122223120002)
【分类号】：S567.219

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