薰衣草芳樟醇合酶基因的克隆、表达及酶活性检测

Cloning, Expression and Enzyme Activity Detection of Linalool Synthase Gene in Lavender

从高油薰衣草品种"杂花"中克隆得到芳樟醇合酶基因LIS1和LIS2,并对其进行了生物信息学和表达模式分析、原核表达和酶活性检测,以低油品种"法国蓝"为对照。结果表明,LIS1基因编码602个氨基酸,LIS2基因编码564个氨基酸。LIS1蛋白与阔叶薰衣草、一串红的芳樟醇合酶亲缘关系相近,LIS2蛋白与狭叶薰衣草、杂薰衣草的芳樟醇合酶亲缘关系相近。LIS1基因在"杂花"花器官半开期、盛开期和衰败期的表达量均高于"法国蓝";LIS1基因在"杂花"花萼中的表达量最高,且仅在"法国蓝"雄蕊中少量表达。LIS2基因在"杂花"花器官不同组织和不同发育时期表达量均低于其在"法国蓝"中表达量,该基因在"杂花"花器官衰败期和"法国蓝"花器官半开期表达量最高,在2个品种花萼中高表达。LIS2基因在2个品种不同发育时期及组织中的表达量均明显高于LIS1基因。LIS1和LIS2重组蛋白具有单萜合酶活性,且均参与合成芳樟醇。

In this study, the LIS1 and LIS2 genes of lavender were cloned from the oil-rich lavender variety ‘Zahua’, and bioinformatics analysis, expression pattern analysis and prokaryotic expression and enzyme activity detection with the control of ‘French bule’ were carried out. The results showed that LIS1 gene could encode 602 amino acids and LIS2 gene could encode 564 amino acids. LIS1 protein was closely related to the linalool synthase in broadleaf Lavandula latifolia and Salvia splendens. The LIS2 protein was closely related to the linalool synthase in Lavandula angustifolia and Lavandula x intermedia. The expression level of the LIS1 gene in half-opening stage,full blooming stage and senescence stage of flower organs of ‘Zahua’ were higher than that of ‘French blue’ at the same stages. The expression of the LIS1 gene was the highest in calyx of ‘Zahua’, and only slight expression in the stamen of ‘French blue’. The expression level of the LIS2 gene in different tissues and different development stages of ‘Zahua’ flower organs were lower than that of ‘French blue’ in the same tissues and stages. The LIS2 gene had the highest expression in ‘Zahua’ flower organs at senescence stage and in ‘French blue’ flower organs at the halfopening stage, and was strongly expressed in the calyx of the two varieties. The LIS2 gene expression was significantly higher than that of the LIS1 gene in different development stages and tissues of the two varieties flower organs. The recombinant proteins LIS1 and LIS2 had monoterpene synthase activity and were involved in the synthesis of linalool.