蕙兰CfMADS1基因的克隆及其时空表达特性  

Cloning and spatiotemporal expression of CfMADS1 gene in orchid(Cymbidium faberi)

作　　者：田云芳[1,2] 袁秀云[3] 蒋素华[3] 马杰[3] 苏金乐[2] 崔波[3] 

机构地区：[1]郑州师范学院生命科学学院,河南郑州450000 [2]河南农业大学林学院,河南郑州450000 [3]郑州师范学院生物研究所,河南郑州450000

出　　处：《西北农林科技大学学报（自然科学版）》2015年第7期143-149,154,共8页Journal of Northwest A&F University(Natural Science Edition)

基　　金：河南省科技攻关项目(092102110128);河南省教育厅科学技术研究重点项目(14A22005)

摘　　要：【目的】对蕙兰CfMADS1基因cDNA全长进行克隆和时空表达特性研究,为研究成花相关基因的功能提供参考。【方法】以蕙兰萼片cDNA为模板,利用反转录RT-PCR和RACE的方法,克隆获得CfMADS1全长cDNA序列,并对其进行生物信息学分析。以蕙兰各时期的器官组织为材料,利用荧光定量PCR进行该基因的时空表达分析。【结果】成功克隆了CfMADS1基因(GenBank登录号为KC148540),其cDNA序列全长1 061bp,开放阅读框长744bp,编码247个氨基酸,分子式为C1 244H2 040N370O377S9。其编码的氨基酸序列与AP1/FUL亚家族中金钗石斛MADS1具有较高的同源性(85.43%),与AP1/FUL转录因子亚家族中的蛋白聚为一类。该基因编码蛋白没有明显的信号肽和跨膜域,极可能位于细胞核内,具有MADS保守域(1-61aa)和相对保守的K区(87-178aa),而且在C末端具有AP1/FUL特征基序(LPPWML),二级结构中α-螺旋所占比例较高(55.63%),三级结构与月季、水稻和水仙MADS1蛋白结构非常相似。该基因在盛花期叶中表达最强烈,花蕾期叶、葶、蕾及营养期叶、盛花期葶中表达较为强烈,子房和营养期根中表达较弱,其余器官组织中表达痕量。【结论】从蕙兰萼片中克隆得到了CfMADS1cDNA全长序列,CfMADS1与蕙兰的成花诱导、花发育以及果实形成有关。[Objective] This study cloned a full-length APETALA1/FRUITFULL-Iike （AP1/FUL- like） MADS box cDNA from Cymbidium faberi and analyzed its spatiotemporal expression to provide a reference for studying function of flower related genes. [Method] CfMADS1 gene was isolated from sepals of C. faberi by RT-PCR and RACE. Then spatiotemporal expression was detected by real time PCR. [Result] CfMADS1 gene was cloned （GenBank number： KC148540）. The full length of cDNA sequence is 1 061 bp, with open reading frame of 744 bp, encoding 247 amino acids. The molecular formula is C1 244 H2 040 N370 O377S9. The deduced amino acid sequence of CfMADS1 shared 85.43 % homology with a member of the AP1/FUL-like group of MADS box genes （Dendrobium nobile MADS1）. CfMADS1 belonged to the AP1/ FUL transcription factor subfamily. The deduced protein has no obvious signal peptide and transmembrane domain,and probably located in the nucleus. It had a MADS domain （1-61aa）,a relatively conservative K region（87- 178aa）and a C terminal conserved motif LPPWML. The secondary structure of CfMADS1 mainly consisted of alpha helices （55.63%）,and the three-dimensional structure of the protein was similar to that of homologues in Roza hybrida ,Oryza sativa and Narcissus tazetta. Real-time quantitative PCR （qRT-PCR） results revealed low levels of its mRNA in ovaries and roots during vegetative period,and in other tissues. Highest levels of CfMADS1 expression were observed in leaves at full-blossom stage followed by pedicels at reproductive period, leaves at vegetative period, and leaves, pedicels, and buds at bud stage. [Conclusion] A full-length AP1/FUL-like MADS box cDNA was cloned from C. faberi sepals and CfMADS1 involved in floral induction,floral development and fruit formation.

关 键 词：蕙兰 AP1/FUL-like基因 时空表达 

分 类 号：Q786[生物学—分子生物学]