珙桐DiZF-CCCH1基因的克隆及表达分析  

Cloning and expresssion analysis of DiZF-CCCH1 gene in Davidia involucrata

作　　者：纪小敏 李建 李萌[1] 曹福祥 董旭杰[1] JI Xiaomin;LI Jian;LI Meng;CAO Fuxiang;DONG Xujie(College of Life Science and Technology,Central South University of Forestry&Technology,Changsha 410004,Hunan,China;Horticulture&Landscape College,Hunan Agricultural University,Changsha 410128,Hunan,China)

机构地区：[1]中南林业科技大学生命科学与技术学院,湖南长沙410004 [2]湖南农业大学园艺园林学院,湖南长沙410128

出　　处：《中南林业科技大学学报》2020年第5期154-161,共8页Journal of Central South University of Forestry & Technology

基　　金：湖南省重点研发计划项目(2017NK2252);湖南省教育厅科学研究项目(18C0268)。

摘　　要：【目的】珙桐Davidia involucrata有性生殖能力弱是限制其自然更新的主要原因。为研究珙桐种子发育的分子机制,本课题组前期完成了珙桐种子转录组分析,筛选到1个编号为c37849.graph_c0的转录本,其在正常种子中高量表达,而在发育异常的种子中表达量很低,差异表达显著,因此将其作为研究目标。【方法】使用PCR克隆该转录本的全长序列,并利用在线分析工具对其编码氨基酸的序列特征、蛋白结构和系统进化等进行生物信息学分析预测,采用qRT-PCR检测目标基因在珙桐不同组织部位以及种子不同发育阶段中的表达情况。【结果】序列分析确定目标基因为一个编码CCCH型锌指蛋白转录因子的基因,将其命名为DiZF-CCCH1,该基因开放阅读框全长为711 bp,编码236个氨基酸,无内含子,相对分子量26.74 kDa,为不稳定蛋白,其氨基酸序列含有植物特有的RR-TZF结构域,与来源于甜橙Citrus sinensis的CCCH蛋白同源性最高。基因表达模式分析显示,DiZF-CCCH1在种子发育前期的表达量最高,后期表达量降低,在其他组织中仅有微量表达,推测其可能在种子发育过程中发挥重要作用。【结论】对珙桐DiZF-CCCH1基因进行了克隆与初步生物信息学分析,发现其编码的蛋白属于RR-TZF型锌指蛋白,且DiZF-CCCH1可能是珙桐种子发育过程中的重要调控基因,为进一步探索该基因在珙桐生殖发育过程中的功能奠定基础。【Objective】The sexual reproduction of dove tree(Davidia involucrata) is the main determinant for the restriction of its natural regeneration. In order to reveal the molecular mechanism of seed development in Davidia, we performed the transcriptome analysis of Davidia seeds, and identified a transcript with the code of c37849. graph_c0, which was highly expressed in normal seeds, but very low in abnormal seeds, so it was taken as the research target.【Method】The full-length sequence of the transcript was cloned by PCR, and the sequence characteristics, protein structure and phylogeny of amino acids encoded by the transcript were predicted by online analysis tools. QRT PCR was used to detect the expression of the target gene in different tissues and seeds of Davidia involucrata.【Result】Sequence analysis identified the target gene as a gene encoding a CCCH-type zinc finger protein transcription factor, which was named DiZF-CCCH1. The open reading frame of this gene is 711 bp in length, encoding 236 amino acids, no intron, and the relative molecular weight is 26.74. kDa, an unstable protein, has an amino acid sequence containing a plant-specific RR-TZF domain, and has the highest homology with a CCCH protein derived from Citrus sinensis. Gene expression pattern analysis showed that DiZF-CCCH1 had the highest expression level in the early stage of seed development, and decreased the expression level in the later stage. It was only slightly expressed in other tissues, suggesting that it may play an important role in seed development.【Conclusion】The DiZF-CCCH1 gene was cloned and preliminary bioinformatics analysis. It was found that the encoded protein belongs to RR-TZF type zinc finger protein, and DiZF-CCCH1 may be an important regulatory gene in the development of Davidia involucrata. To further explore the role of this gene in the reproductive development process of Davidia involucrata.

关 键 词：珙桐 锌指蛋白转录因子 DiZF-CCCH1 种子 

分 类 号：S792.99[农业科学—林木遗传育种]