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本研究采用RT-PCR技术从青花菜自交系‘WN12-95B’中克隆到谷胱甘肽S-转移酶基因。序列分析表明,青花菜GST基因全长690 bp,ORF长度为675 bp,推导编码蛋白含有224个氨基酸,相对分子量为26.15 kD,理论pI值为6.56,属于Tau类家族成员。青花菜GST蛋白的二级结构主要以α-螺旋和延伸链为主。通过构建系统进化树发现,GST基因与油菜、芜菁亲缘关系最近。利用荧光定量PCR检测GST基因在青花菜保持系不同组织中的表达量,结果显示GST基因在根、叶、荚中的表达丰度较高,花蕾中表达丰度较低。青花菜Ogu不育系及其保持系不同发育阶段花蕾时空表达特性分析表明,花蕾发育早期(<2 mm)表达量最高,随着发育进程的推移,表达量逐渐下降。同时期不育系的表达量高于保持系。本研究为探讨青花菜GST基因在花粉发育过程中的功能提供一定理论依据。
In this study, glutathione S-transferase gene was cloned from the inbred line ’WN12-95B’ of broccoli by RT-PCR. Sequence analysis showed that the GST gene was 690 bp in length and 675 bp in length. The deduced protein contained 224 amino acids with a relative molecular mass of 26.15 kD and a theoretical pI value of 6.56, belonging to the Tau family of members. The secondary structure of broccoli GST protein is dominated by α-helix and extension chain. Through constructing phylogenetic tree, we found that GST gene has the closest genetic relationship with rape and turnip. The expression of GST gene in different tissues of broccoli maintainer lines was detected by fluorescence quantitative PCR. The results showed that GST gene expressed higher in roots, leaves and pods and lower in flower buds. Analysis of the temporal and spatial expression of buds in Ogu CMS lines and their maintainer lines at different developmental stages showed that the expression of Ogu and its maintainer lines was the highest in early bud development (<2 mm), and decreased gradually with the progress of development. At the same time, the expression of CMS was higher than that of maintainer line. This study provides some theoretical basis for exploring the function of GST gene in pollen development.