烟草花青素苷转运相关基因NtAN9的分离与表达分析

摘要

To study anthocyanin transport ,we isolated a full-length gene NtA N9 (GenBank Accession No . KX356542) by in silicon cloning and RT-PCR method from Nicotiana tabacum .The NtA N9 contains a 690 bp open reading frame (ORF) encoding 229 amino acid residues ,which belongs to a putative phi class glutathiones-transferase(GST)family.Itsgenomicstructurecontains3exonsand2introns.Multiplese-quence alignment showed that NtAN9 has highest identities (88% ) with petunia PhAN9 that is an antho-cyanin transport-related GST .Phylogenetic analysis showed that NtA N9 was clustered into an anthocya-nin transport-related GST group ,and was an ortholog of PhA N11 .The anthocyanin content analysis and real-time quantitation PCR (qRT-PCR) showed that NtA N9 was expressed in different flower develop-mental stages ,with highest level in stage Ⅲ ,but almost no expression in root ,stem and leaf w hich are al-most no anthocyanin .These results suggest that the NtA N9 function is similar with PhA N9 and might be related to the transfer and accumulation of anthocyanin in tobacco plants .%为研究花青素苷的转运，利用电子克隆和RT-PCR方法，从普通烟草（Nicotiana tabacum）花中分离了1个编码谷胱甘肽转移酶（GST）基因，命名为 NtAN9（GenBank登录号KX356542）。 NtAN9包含一个690 bp的开放阅读框，编码229个氨基酸残基，属于phi型GST 。 N tA N9基因组结构由3个外显子和2个内含子组成。多序列比对分析表明，NtAN9与矮牵牛（Petunia hybrida）花青素苷转运相关的GST 基因 PhAN9具有88％的一致性。系统进化分析显示，N tA N9与花青素苷转运相关的GST 基因聚为一支，是 PhA N9的直系同源基因。定量 PCR分析表明，N tA N9基因在含有花青素苷的四个花发育时期中均有表达，其中在开花前的第Ⅲ期（2 cm ＜花芽＜4 cm ）表达丰度达到最高，而在不含花青素苷的根、茎和叶中不表达。由此推测，分离得到的 N tA N9可能具有类似PhA N9的功能，与烟草花青素苷的转运与积累相关。 N tA N9基因的分离与表达分析，为进一步研究烟草花青素苷的转运奠定了基础。