茶树开花相关基因家族的克隆及CsMFT基因的可变剪切分析

Cloning of Flowering-Related Genes and Alternative Splicing Analysis of CsMFT in Tea Plant

开花是植物从营养生长到生殖生长转变的关键过程,PEBP(phosphatidylethanolamine-binding protein)蛋白家族在植物开花过程中发挥重要调控作用。该研究分别采用信息学分析及RT-PCR方法,对茶树CsPEBP基因家族进行了鉴定、克隆和表达分析。结果表明:(1)成功克隆获得5个PEBP基因家族成员,并分别命名为CsATC、CsMFT、CsBFT、CsFT和CsTFL1,其长度为519~525 bp,分别编码172~174个氨基酸残基,定位于5条不同染色体。(2)结构分析显示,该蛋白家族不同成员氨基酸序列的相似性高达72.7%,含39.88%~42.28%的自由卷曲,分属于3个亚家族,其亲缘关系与杨树最近。(3)亚细胞定位分析显示,CsATC、CsMFT、CsBFT定位于细胞质,CsFT定位于细胞核,CsTFL1定位于细胞质和细胞核。(4)转录组和荧光定量PCR分析显示,CsMFT基因在茶树不同组织部位和不同非生物胁迫响应下的表达量均高于其他基因;CsFT、CsATC、CsMFT基因在茶树花半开时的表达量最高。(5)启动子元件分析显示,该基因家族的启动子中含有大量的光响应元件和激素响应元件。(6)CsMFT基因存在可变剪切,有525 bp和689 bp两个不同长度的转录本。研究推测,该研究所克隆的5个茶树CsPEBP家族成员均参与了调控茶树的开花过程和茶树对多种逆境的响应,为茶树开花调控相关研究奠定了基础。

Flowering is the key process in the transformation from vegetative growth to reproductive growth in plants.The PEBP(phosphatidylethanolamine-binding protein)proteins played important roles in this process.In this study,the CsPEBP genes were identified,cloned and analyzed in tea plant.The results showed that:(1)five members of the PEBP gene family were identified and cloned from the genome of tea plant,named CsATC,CsMFT,CsBFT,CsFT and CsTFL1,respectively,and their lengths were 519-525 bp,encoding 172-174 amino acid residues,located in 5 different chromosomes,respectively.(2)The structural analysis showed that the homology of amino acid sequence of this protein family is as high as 72.7%,containing 39.88%-42.28%of free curl,belonging to 3 subfamilies,and the genetic distance of these proteins between tea plant and poplar was the closest.(3)The result of subcellular location showed that CsATC,CsMFT and CsBFT were located in cytoplasm,CsTFL1 was located in nucleus,CsFT was both located in cytoplasm and nucleus.(4)Transcriptome and realtime-PCR showed that CsMFT gene had the higher expression in different tissues and under different abiotic stresses than other genes in tea plant;The expression of CsFT,CsATC and CsMFT genes were the highest in the stage of half-opened flower.(5)Promoter analysis showed that a large number of light-responsive elements and hormone-responsive elements were found in the promoters of CsPEBP genes.(6)CsMFT gene had two transcripts with different lengths of 525 bp and 689 bp.In summary,we cloned five CsPEBPs genes and speculated that they not only participate in the regulation of flowering,but also in the response to various adversities in tea plants,which provided a theoretical basis for the research on the flowering regulation of tea plants.