茶树△12-脂肪酸去饱和酶基因FAD2和FAD6的克隆与表达分析

摘要/Abstract

摘要： 本研究在茶树转录组测序的基础上,以铁观音茶树的芽叶为材料,采用RT-PCR技术,克隆了茶树不饱和脂肪酸合成途径中的关键限速酶—△12-FAD（△12-脂肪酸去饱和酶）基因的包含完整ORF的cDNA序列（CsFAD2和CsFAD6）。生物信息学分析结果表明,CsFAD2的全长为1β184βbp,其开放阅读框（ORF）长度1β149βbp,编码382个氨基酸,定位于内质网上,其氨基酸序列与油茶FAD2的同源性最高达97%;CsFAD6的全长为1β425βbp,其ORF长度为1β311βbp,编码436个氨基酸,定位于叶绿体上,其氨基酸序列与葡萄FAD6同源性达81%。荧光定量PCR结果表明,铁观音茶树幼苗在4℃低温胁迫处理72βh过程中,这两个基因的表达均受低温的诱导,其表达量随着处理时间的延长而升高,在处理48βh时,表达量水平最高;在100βg·L^-1的PEG胁迫处理12βh过程中,这两个基因的表达均受PEG胁迫处理的诱导;在ABA（100βμmol·L^-1）胁迫处理72βh过程中,在处理6~24βh期间,CsFAD2的表达量显著升高,而CsFAD6的表达不受ABA处理的影响,CsFAD6的表达量在处理72βh时显著降低;在NaCl（250βmmol·L^-1）胁迫72βh过程中,CsFAD2的表达量全程降低,而CsFAD6在处理24~72βh期间表达量显著升高。

关键词: 茶树, △12-脂肪酸去饱和酶;, 非生物胁迫, 基因表达

Abstract: Based on the transcriptome database of tea plants, the cDNAs ( CsFAD2 and CsFAD6) including full ORFs of the key rate-limiting enzyme-FAD (△12-fatty acid desaturase) in the unsaturated fatty acid synthesis pathway were cloned from the buds and leaves of tea cultivar Tieguanyin by RT-PCR .The full length of cDNA of CsFAD2 was 1 184 bp, which contained a 1β149βbp ORF encoding 382 amino acids with 97% homologous to the CoFAD2. Subcellular localization prediction showed that CsFAD2 was localized to the endoplasmic reticulum. The full length of cDNA of CsFAD6 was 1β425βbp and contained a 1β311βbp ORF encoding 436 amino acids, which was 81% homologous to VvFAD6. It was predicted to be located on the chloroplast. Quantitative PCR analysis showed that the expression of CsFAD2 and CsFAD6 were induced by cold stress (4℃), which had the highest expression levels at 48βh. Similarly, CsFAD2 and CsFAD6 were induced by 100βg·L^-1 PEG treatment for 12βh. The expression of CsFAD2 was significantly up-regulated under the treatment of ABA(100βμmol·L^-1) for 6-24βh. While the expression of CsFAD6 was not affected by ABA during this period and dramatically down-regulated at 72βh. The expression of CsFAD2 was repressed under NaCl (250βmmol·L^-1) treatment. Inversely;|the expression of CsFAD6 was significantly induced by NaCl treatment from 24 to 72βh.

Key words: Camellia sinensis, △12-fatty acid desaturase enzyme, abiotic stresses, gene expression

中图分类号:

陈丹, 俞滢, 岳川, 王鹏杰, 陈静, 陈桂信, 叶乃兴. 茶树△12-脂肪酸去饱和酶基因FAD2和FAD6的克隆与表达分析[J]. 茶叶科学, 2017, 37(6): 541-550.

CHEN Dan, YU Ying, YUE Chuan, WANG Pengjie, CHEN Jing, CHEN Guixin, YE Naixing. Cloning and Expression Analysis of △12-fatty Acid Desaturase in Tea Plants[J]. Journal of Tea Science, 2017, 37(6): 541-550.

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