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目的克隆出浙贝母法呢基焦磷酸合酶(Farnesyl pyrophosphate synthase,FPS)基因,并对其表达量与生物碱积累量之间的相关性进行分析,以探究FPS基因在浙贝母生物碱合成代谢过程中的调控作用。方法本研究采用RACE技术克隆出浙贝母FPS基因的全长序列;以浙贝母狭叶、宽叶和新梅园3个品种6个组织及10个不同发育时期的新鳞茎为材料,采用RT-qPCR和HPLC-ELSD技术分别测定了FPS基因的表达量与生物碱(浙贝甲素和浙贝乙素)的量,并对其进行相关性分析。结果 FPS基因cDNA序列全长1 629 bp,开放阅读框为1 056 bp,编码351个氨基酸;3个不同浙贝母品种中FPS基因组织特异性表达趋势相同,花中的表达量最高,其次为新鳞茎;不同发育时期新鳞茎FPS基因表达量与生物碱量相关性分析显示,在宽叶和新梅园品种中,FPS基因表达量与浙贝甲素、浙贝乙素量均具有明显的正相关性(相关系数分别为0.289、0.613,0.427、0.622),但在狭叶品种中,该相关性相对较低(0.054、0.236);狭叶、宽叶和新梅园3个品种不同组织中FPS基因表达量与生物碱量具有正相关性(相关系数分别为0.057、0.476,0.085、0.495,0.375、0.432)。结论本研究成功克隆出浙贝母FPS基因的全长序列,FPS基因参与了浙贝母生物碱的合成代谢调控。
Objective To clone the FPS gene of Fritillaria thunbergii and analyze the correlation between the expression of FPS gene and alkaloid accumulation, Regulatory role in the process of anabolism. METHODS: The full-length FPS gene of Fritillaria cirrhosa was cloned by RACE technique. Six Fusarium oxysporum f. Sp., Ficus macrocephala and Xinmeiyuan 6 cultivars and 10 new bulbs at different developmental stages were used as materials. RT-qPCR and HPLC-ELSD techniques were used to determine the amount of FPS gene expression and the amount of alkaloids (Zhe Pei A and Zhe B), and their correlation analysis. Results The sequence of FPS cDNA was 1 629 bp in length and 1 056 bp in open reading frame, encoding 351 amino acids. The FPS gene expression in 3 different Fritillaria species was the same with the highest expression in flowers, followed by The results showed that the expression of FPS gene was positively correlated with the amount of Fupivan and Zhebukin in the broad-leaved and Xinmei orchard cultivars The correlation coefficient was 0.289,0.613,0.427,0.622, respectively. However, in the narrow-leaf variety, the correlation was relatively low (0.054,0.236). In the three varieties of narrow-leaved, broad- There was a positive correlation between FPS gene expression and alkaloid amount (correlation coefficients were 0.057, 0.476, 0.085, 0.495, 0.375, 0.432, respectively). Conclusion The full-length FPS gene of Fritillaria cirrhosa was successfully cloned in this study. FPS gene is involved in the synthesis and metabolism of Fritillaria cirrhosa.