枸杞果糖激酶基因LbFRK7的克隆及表达分析
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国家自然科学基金(31360191,31660220);


Cloning and Expression Analysis of Fructokinase Gene (LbFRK7) from Wolfberry (Lycium barbarum Linn.)
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    摘要:

    该研究以‘宁杞1号’枸杞果实为材料,基于转录组测序TR28373|c0_g1序列,利用RTPCR技术, 克隆出枸杞果糖激酶基因LbFRK7的全长序列为1 060 bp,其中,ORF开放阅读框为1 044 bp,编码有348个氨基酸,蛋白质分子量为37.44 kD,理论等电点5.05; LbFRK7编码的氨基酸序列包含有pfkB碳水化合物激酶家族高度保守的3个特异性区域, 2个底物识别位点,4个ATP结合位点;LbFRK7与烟草和辣椒的FRP7基因序列相似性较高,达到90%;利用实时荧光定量技术分析发现,不同组织中LbFRK7基因均有表达,且果实中的表达量最高,根中最低;随着果实的发育,果实中LbFRK7基因的表达量呈先升后降的变化趋势,并于开花后15 d达到最高。在果实发育前期,LbFRK7基因的表达量与果糖含量的变化趋势相同,但在果实发育中期和后期,LbFRK7基因的表达量与果糖含量的变化趋势相反。相关分析结果显示,LbFRK7表达量与果糖和蔗糖含量的相关系数分别为-0.326和-0.339,但均未达到显著水平。研究表明,LbFRK7基因在枸杞果实发育过程中对果糖转化起到一定作用,特别是在果实成熟过程中对果糖含量的升高具有重要的作用。该研究结果为进一步探讨枸杞LbFRK7的功能及果糖代谢奠定了基础。

    Abstract:

    In this study, the fulllength sequence of fructokinase gene (LbFRK7) was obtained by using RTPCR from wolfberry (Lycium barbarum Linn.) ‘Ningqi No.1’ fruit based on TR28373|c0_g1 transcriptome sequencing. The fulllength sequence of LbFRK7 was 1 060 bp (ORF length 1 044 bp)and encoding a polypeptide of 348 amino acids. The relative molecular weight was 37.44 kD and the theoretical isoelectric point (pI) was 5.05. The amino acid sequence of LbFRK7 contained three highly conserved specific regions interent to the pfkB family of carbohydrate kinases, two substrates recognition locus and four ATP binding locus. Homologous genes comparison indicated that LbFRK7 had more high similarity with tobacco and pepper fructokinase genes(90% homology). The change of LbFRK7 expression was founded in different plant tissues by quantitative realtime PCR. LbFRK7 expressed in the highest in fruits and in the lowest in roots. LbFRK7 expression of fruits showed a trend on firstly rising and then falling during fruit ripening, and reached on the highest level at 15 days after full bloom. LbFRK7 expression had a similar trend with fructose content at the early stage of fruit development, while there was an opposite change between LbFRK7 expression and fructose content at the mid and late stages. LbFRK7 expression was not significantly correlation with fructose and sucrose contents by correlation analysis (r1=-0.326, r2=-0.339). Therefore, LbFRK7 played a role on converting fructose during wolfberry fruit development, particularly on increasing of fructose content during the process of fruit ripening. This study provided a reference basis on researching into function of LbFRK7 and metabolism of fructose.

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赵建华,尹 跃,李浩霞,等.枸杞果糖激酶基因LbFRK7的克隆及表达分析[J].西北植物学报,2018,38(5):816-822

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  • 在线发布日期: 2018-06-14
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