发芽苦荞GABA-T基因克隆及序列分析

Cloning and analysis of the GABA-T gene from germinated tartary buckwheat

γ-氨基丁酸转氨酶(γ-aminobutyric acid transaminase GABA-T,EC2.6.1.19)是植物GABA转化途径中的第一步关键酶,与γ-氨基丁酸GABA的积累具有一定的关系.文中将苦荞进行发芽处理提取其总RNA,利用反转录聚合酶链式反应(reverse transcription polymerase chain reaction,RT-PCR)、巢氏PCR(nested PCR)与3′-cDNA末端快速扩增技术(rapid amplification of cDNA end,RACE)成功获得发芽苦荞GABA-T全长基因.序列分析表明,该基因序列全长1556 bp,含有一个编码448个氨基酸的开放阅读框(open reading frame,ORF),其氨基酸序列与菠菜全长GABA-T氨基酸序列和欧洲栓皮栎树全长GABA-T氨基酸序列的相似度分别为87.05%和86.83%.基因序列的进化树分析结果表明,发芽苦荞GABA-T基因与甜菜、藜麦和菠菜来源相关基因的亲缘性最近.

GABA transaminase GABA-T(EC 2.6.1.19)is the first key enzyme in the plant GABA transformation pathway,and is considered to have a certain relationship with the accumulation of GABA.In this study,the total RNA was extracted from germinated tartary buckwheat and the GABA-T gene was successfully cloned based on reverse transcription polymerase chain reaction(RT-PCR),nested PCR and rapid amplification of cDNA 3′-end(RACE)technologies.Sequence analysis showed that it was 1556 bp in length and contained an open reading frame(ORF)encoding 448 amino acids.Its amino acid sequence has 87.05%and 86.83%similarity to the full-length GABA-T amino acid sequence of Spinacia oleracea and Eucalyptus urophylla,respectively.Phylogenetic tree analysis showed that it has the closest genetic relationship with sugar beet,buckwheat and spinach.