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为揭示牻牛儿基牻牛儿基焦磷酸合成酶(GGPPS)小亚基在烟草(Nicotiana tabacum)中的生理功能,采用电子克隆的方法,结合RT-PCR和SMART RACE技术,从烟草中克隆到1个牛儿基牛儿基焦磷酸合成酶小亚基基因的cDNA序列,命名为NtGGPPS5(GenBank登陆号:KF316932)。该基因全长1320 bp,编码332个氨基酸,与番茄(Solanum lycopersicum,XP_004246572)及其野生种(Solanum pennellii,ADZ24721)的GGPPS小亚基的氨基酸序列一致性均为92%,具有一个特征性的异戊二烯合成酶保守的天门冬氨酸富集区。进化分析表明,植物GGPPS分为大亚基和小亚基两个分支,而且NtGGPPS5属于小亚基。实时荧光定量PCR试验表明,NtGGPPS5基因在烟草根、茎、叶和芽中均有表达,表达量为芽>叶>茎>根。
In order to reveal the physiological function of geranylgeranyl diphosphate synthase (GGPPS) small subunit in tobacco (Nicotiana tabacum), we cloned it from tobacco using electron cloning method combined with RT-PCR and SMART RACE To 1 cDNA of the small subunit gene of geranylgeranyl pyrophosphate synthase named NtGGPPS5 (GenBank accession number: KF316932). The full length of this gene was 1320 bp, encoding a protein of 332 amino acids. The deduced amino acid sequence was 92% identical to the GGPPS small subunit of Solanum lycopersicum (XP_004246572) and its wild species (Solanum pennellii, ADZ24721), with a characteristic Isoprene synthase conserved aspartate-rich region. Phylogenetic analysis showed that plant GGPPS is divided into two subunits: big subunit and small subunit, and NtGGPPS5 belongs to small subunit. Real-time PCR showed that the NtGGPPS5 gene was expressed in roots, stems, leaves and buds, and the expression levels were bud> leaf> stem> root.