作物学报2016，Vol.42Issue(1)：58-69,12.DOI:10.3724/SP.J.1006.2016.00058
茶树生长素外运载体基因CsPIN3的克隆与表达分析
Cloning and Expression Analysis of Auxin Efflux Carrier Gene CsPIN3 in Tea Plant (Camellia sinensis)
摘要
Abstract
On the basis of previous transcriptome study on tea plant cold acclimatization, we obtained a PIN homology gene named CsPIN3 and cloned its full-length cDNA sequence by reverse transcription-PCR (RT-PCR) combining with rapid amplifi-cation of cDNA ends (RACE). The full length cDNA of CsPIN3 is 2654 bp (GenBank accession No. KP896474) and contained a 1926 bp open reading frame (ORF) encoding 641 amino acid residues. Bioinformatic analyses showed that CsPIN3 is not a secre-tory protein and had a molecular weight of 70.15 kD, atheoretical isoelectric point of 8.42. Subcellular localization prediction showed that CsPIN3 is a typical membrane protein mainly located in plasmalemma and then in endoplasmic reticulum. Moreover, amino acid sequence analysis indicated that CsPIN3 protein contained hydrophobic regions in both ends and hydrophilic regions in the middle. Similar to PIN protein in rice, the hydrophobic regions of CsPIN3 consisted of several transmembrane helixes, among which five was in N motif and four in C motif. The hydrophilic regions of CsPIN3 had two unstable domains, several o-glycosylation sites, several phosphorylation sites like TPRXS (N/S) motif (a PID/PINOID phosphorylation site) and a well characterized conserved inner motif NPNXY regulating the endocytosis of PIN. Comparison of sequences similarity showed that the amino acid sequence coded by CsPIN3 had more than 80%similarity with reported PINs of Populus trichocarpa, Vitis vinifera, Citrus sinensis, Nicotiana tomentosiformis, Solanum lycopersicum, Solanum tuberosum, and Sesamum indicum. Phylogenetic tree analysis showed that CsPIN3 had the closest genetic relationship with Solanaceae and the highest identity with AtPIN3 of Arabi-dopsis thaliana PIN proteins. The CsPIN3 gene differentially expressed in different tea plant tissues, and transcript abundance in flower was much higher than that in leaf, stem and root. In addition, we analyzed the expression of CsPIN3 by qRT-PCR during the different phases of bud dormancy formation and break, and the results indicated that in cultivar Longjing 43, the expression level of CsPIN3 at growth phase was higher than that at dormant phase (from initial dormant stage to expanding stage) and an obvious expression jump was detected at bud sprouting stage. These results demonstrated that CsPIN3 might be associated with the regulation of tea plant bud dormancy formation and break.关键词
茶树/休眠/生长素外运载体基因/表达分析Key words
Tea plant (Camellia sinensis)/Dormancy/Auxin efflux carrier/Expression analysis引用本文复制引用
王博,曹红利,黄玉婷,胡玉荣,钱文俊,郝心愿,王璐,杨亚军,王新超..茶树生长素外运载体基因CsPIN3的克隆与表达分析[J].作物学报,2016,42(1):58-69,12.基金项目
本研究由国家自然科学基金项目(31370690),国家现代农业产业技术体系建设专项(CARS-23)和中国农业科学院科技创新工程(CAAS-ASTIP-2014-TRICAAS)资助。This study was supported by the National Natural Science Foundation of China (31370690), the Special Program of Modern Agro-industry Technology System (CARS-23) and the Agricultural Science and Technology Innovation Program of Chinese Academy of Agricultural Sciences (CAAS-ASTIP-2014-TRICAAS) （31370690）
