龙眼体胚发生过程中DCL基因的分子特性及表达分析  

Molecular Characterization and Expression Patterns of the DCL Gene Family during Somatic Embryogenesis in Dimocarpus longan Lour.

作　　者：陈晓慧[1] 白玉[1] 陈旭[1] 李汉生[1] 林玉玲[1] 赖钟雄[1] CHEN Xiaohui;BAI Yu;CHEN Xu;LI Hansheng;LIN Yuling;LAI Zhongxiong(Institute of Horticultural Biotechnology, Fujian Agriculture and Forestry University, Fuzhou 350002, China)

机构地区：[1]福建农林大学园艺植物生物工程研究所,福州350002

出　　处：《西北植物学报》2017年第11期2120-2129,共10页Acta Botanica Boreali-Occidentalia Sinica

基　　金：国家自然科学基金(31572088;31672127);福建省重大专项(2015NZ0002-1);福建省自然科学基金杰出青年项目(2015J06004)

摘　　要：从龙眼转录组unigene序列筛选获得龙眼DCL基因(命名为DlDCL)全长序列,并结合生物信息学及实时荧光定量等方法,对龙眼DCL基因进行研究,以明确DlDCLs在龙眼体胚、不同生长组织部位中的表达规律及其对激素和光质应答反应,为进一步研究龙眼体胚过程中DlDCLs基因的调控研究奠定基础。结果表明:(1)龙眼转录组数据存在DlDCL1、DlDCL2、DlDCL3和DlDCL4四个DCL家族成员,且基于Unigene的FPKM值发现不同基因在体胚发生阶段具有差异表达。(2)生物信息学分析发现,DlDCLs成员间基本理化性质较为类似,均为亲水性不稳定蛋白、不含信号肽、可进行跨膜运动,但也存在一定差异,如DlDCL2为碱性蛋白,而其他3个成员为酸性蛋白;亚细胞定位预测显示,DlDCLs均定位于细胞核中,但DlDCL2也存在定位于叶绿体中;对DCL蛋白结构域预测显示,DCL是高度保守的蛋白。(3)系统进化树分析显示,不同物种的DCL分为4个分支,同源的DCL蛋白都聚为一类,且DlDCLs与柑橘DCL亲缘关系更为接近。(4)实时荧光定量PCR分析表明,DlDCLs在龙眼非胚性愈伤组织和体胚发生过程中的表达模式差异较大,但DlDCLs在愈伤组织阶段均有较高的表达量,推测DlDCLs在体胚发生过程可能具有功能的独立和协作。DlDCL1和DlDCL2在叶片、花器官等组织部位中的相对表达量较高,暗示DlDCL1和DlDCL2可能参与到光合作用和花器官的发育;DlDCLs还受2,4-D、MeJA、SA激素和光质诱导,表明DlDCLs可能参与激素和光质调控。In the present work,we obtained the full length sequence of DlDCL gene through longan transcriptome data and analyzed them by means of bioinformatics and real-time quantitative PCR.The purpose is to clarify the expression pattern of DlDCLs in longan somatic embryos and different tissues,as well as their expression profiles responses to hormones and light treatment.The study is help to lay the foundation for the further research on the regulation of DlDCLs genes during longan somatic embryos.The results showed that:(1)there are four members of DCL family genes,i.e.DlDCL1,DlDCL2,DlDCL3and DlDCL4,and different DlDCLs genes were differentially expressed at somatic embryogenesis stage based on the FPKM value of Unigene.(2)Bioinformatics analysis showed that the physical and chemical properties of the DlDCLs are mostly similar,all of them are hydrophilic and unstable proteins with transmembrane structure and without signal peptide.Meanwhile,there are also some differences:DlDCL2is basic protein while DlDCL1,DlDCL2and DlDCL4are acidic proteins.Subcellular localization prediction showed that DlDCL2was located in nucleus and chloroplast,while the other three members were located in the nucleus.Protein domain prediction showed that DCLs were highly conserved proteins.(3)Phylogenetic tree analysis showed that the genetic relationship of DlDCLs family in longan were close to those from Citrus sinensis.(4)QPCR analysis showed that the expression patterns of different DlDCLs in non-embryogenic callus(NEC)and somatic embryogenesis(SE)varied a lot,but DlDCLs are relatively high in the stage of embryogenic callus(EC),which may indicated that DlDCLs have distinct but cooperative functions in the process of the NEC and during SE.The expression patterns of DlDCLs in different tissues were also investigated.The results revealed that DlDCL1and DlDCL2had the highest expression in the leaf and followed by floral organ tissues,which suggested that DlDCL1and DlDCL2may participate in photosynthesis and floral organs development.Moreover,Dl

关 键 词：龙眼 体胚发生 DCL基因 分子特性 荧光定量PCR 

分 类 号：Q786[生物学—分子生物学] Q789