| 摘要: |
| 【目的】克隆特早熟春性甘蓝型油菜的FCA同源基因（sBnFCA）及其可变剪接体，并分析该基因的表达模式。【方法】根据拟南芥和芸薹属植物FCA基因的DNA序列和cDNA序列设计引物，用PCR和RT-PCR技术克隆特早熟春性甘蓝型油菜“86号”的FCA同源基因（sBnFCA）及其可变剪接体，对克隆基因及编码蛋白序列进行生物信息学分析，构建系统进化树，并用qR-PCR技术检测sBnFCA基因在不同发育时期根、茎、叶、茎尖中的表达量。【结果】克隆出了sBnFCA基因的全长序列（8 827 bp），得到了sBnFCA-γ可变剪接体及一个新的可变剪接体（sBnFCA-5），新的可变剪接体在GenBank中的登录号为KJ701579.1。sBnFCA-5剪接体的CDS全长1 986 bp，编码662个氨基酸残基，其推导的氨基酸序列具有2个保守的RRM结构域和1个WW结构域。生物信息学分析显示,sBnFCA-5与已报道甘蓝型油菜BnFCA-γ（AF414188.1）的相似性达99%,与拟南芥FCA-γ的相似性达87%。sBnFCA-5比sBnFCA-γ少了172个碱基序列，是一个跨外显子剪接体。sBnFCA蛋白相对分子质量和理论等电点分别为72.5 ku和9.2。基因表达模式分析显示，sBnFCA-γ和sBnFCA-5在苗期、蕾期、花期的根、茎、叶和茎尖组织中都有表达，sBnFCA-γ在蕾期茎尖和花期叶片中表达量较高，sBnFCA-5在蕾期茎尖和花期叶片中表达量极高。【结论】克隆的sBnFCA-5为甘蓝型油菜sBnFCA基因的一个新的可变剪接体，该基因在春性特早熟甘蓝型油菜开花调控中可能有着重要的调节作用。 |
| 关键词: 春性甘蓝型油菜 基因克隆 FCA同源基因 可变剪接 自主开花途径 |
| DOI： |
| 分类号: |
| 基金项目:国家自然科学基金项目（31360345）；青海省科技厅项目（2013-Z-716）；国家863项目（2011AA10A104）；高层次人才项目（2012-QGC-3）；科技部项目（2013BAD01B05 3） |
|
| A novel alternative splicing isoform of FCA homologue in spring rapeseed |
|
ZHANG Shengping,LUO Yuxiu,DU Dezhi,et al
|
| Abstract: |
| 【Objective】The research cloned the sBnFCA gene and its alternative splicing isoform,and analyze their expressions.【Method】The primers were designed according to the published FCA DNA and cDNA sequences of Arabidopsis and Brassica.The genome DNA and complete open reading frame of FCA homologue gene were cloned from an early-flowering rapeseed “No.86” strain using the method of homologues cloning and reverse transcription PCR (RT-PCR) bioinformatics analysis was conducted for cloned gene and proteins before the phylogenetic tree was constructed and the expression levels at different stages were detected using qRT-PCR.【Result】The DNA sequence of FCA from spring B.napus L.(sBnFCA) was 8 827 bp and the cDNA sequence of novel alternative splicing isoform (sBnFCA-5) was 1 986 bp,encoding a protein of 662 amino acid residues with a predicted molecular weight of 72.5 ku and a theoretical pI of 9.2.Bioinformatics analysis showed that the predicted sBnFCA-5 protein contained two RRM domains and a WW domain.It was showed that sBnFCA-5 had 99% similarity with BnFCA-γ（AF414188.1） and had 87% similarity with A.thaliana FCA-γ.The expression analysis of sBnFCA-5 using qRT-PCR showed that the sBnFCA gene was expressed in roots,stem,leaves,shoot tips and buds at seeding,bud and flowering stages.The relative expression level of sBnFCA-5 was highest in shoot tips at bud stage and in leaves at flowering stage,respectively.【Conclusion】The cloned sBnFCA-5 was a new alternative splicing isoform of sBnFCA gene and it might play an important role in flowering regulation of spring rape. |
| Key words: spring rape (Brassica napus L.) gene cloning FCA homologue alternative splicing autonomous flowering pathway |
