枸杞己糖激酶基因LbHXK的克隆及表达分析
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引用本文:李浩霞,杨 斌,尹 跃,安 巍,王亚军,赵建华.枸杞己糖激酶基因LbHXK的克隆及表达分析[J].西北植物学报,2019,39(6):1009~1015
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作者单位
李浩霞1,杨 斌2,尹 跃2,安 巍2,王亚军2,赵建华2* (1 宁夏农林科学院 荒漠化治理研究所银川7500022 宁夏农林科学院 枸杞工程技术研究所/国家枸杞工程技术研究中心银川 750002) 
基金项目:国家自然科学基金(31660220);
中文摘要:该研究以宁夏枸杞‘宁杞1号’果实为材料,利用RT PCR技术, 分离出枸杞己糖激酶基因LbHXK的全长序列(1 494 bp),该基因包含完整开放阅读框ORF,编码498个氨基酸,蛋白质分子量为53.88 kD ,理论等电点5.96; LbHXK基因编码的蛋白具有己糖激酶家族显著的特征,包含一个己糖激酶小亚基 (Gly91~Val228)和一个大亚基结构(Asn229~Asp476),该蛋白三级结构为V型,并且存在一个葡萄糖结合结构域;LbHXK与茄科烟草的同源性最高为93.36%。 实时荧光定量分析发现,LbHXK基因在不同组织均有表达,叶中最高,茎中次之,根中最低,且组织间差异不显著;随着果实发育LbHXK基因表达量呈先升后降的变化趋势,并在色变期(开花后约22 d)达到最高,随后到果实成熟期降至最低,且果实发育前中期显著高于后期。该研究结果为进一步探讨枸杞LbHXK基因的功能奠定了基础。
中文关键词:枸杞  己糖激酶基因  克隆  表达
 
Cloning and Expression Analysis of Hexokinase Gene (LbHXK) from Wolfberry (Lycium barbarum Linn.)
Abstract:In this study,the full length sequence of LbHXK gene was cloned from ‘Ningqi No. 1’ ( Lycium barbarum L.) fruit by RT PCR, and was 1 494 bp. LbHXK gene contained a complete open reading frame (ORF) and encoded a polypeptid of 498 amino acids. The protein molecular weight was 53.88 kD and the theoretical isoelectric point (pI) was 5.96. The LbHXK protein possessed a remarkable feature of the hexokinase family, including a ribosomal small subunit substructure of hexokinase (Gly91 Val228) and a ribosomal big subunit substructure of hexokinase (Asn229 Asp476). The tertiary structure of protein was V type and had a glucose binding domain. Homologous gene comparison indicated that LbHXK had more high similarity with tobacco hexokinase genes (93.36% homology) in Solanaceae. The change of LbHXK expression was founded in different plant tissues by quantitative real time PCR. LbHXK expressed in the highest in leaves and the lowest in roots, and there was no significant difference between different plant tissues. LbHXK expression of fruits showed a trend on first rising and then falling during fruit ripening, among which, reached on the highest level at coloring fruit phase (22 days after full bloom), and reached the lowest at the fruit ripening stage, and was significantly higher at early middle stage than at latter stage of fruit development. This study would providd a foundation on further researching for function of LbHXK.
keywords:Lycium barbarum Linn.  hexokinase gene  clone  expression
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