河南农业大学学报2025，Vol.59Issue(4)：602-613,12.DOI:10.16445/j.cnki.1000-2340.20240912.002
芍药钙依赖性蛋白激酶基因PlCDPK24的鉴定及表达分析
Identification and expression analysis of the calcium-dependent protein kinase gene PlCDPK24 of Paeonia lactiflora
摘要
Abstract
[Objective]The calcium-responsive gene PlCDPK24 was cloned and analysed for expres-sion,laying the foundation for exploring the mechanism by which the CDPK gene regulates the growth of pollen tubes in Paeonia L distant hybridization.[Method]Using the existing RNA-seq database,the calcium-dependent protein family genes in the unaffiliated transcriptome of Paeonia lactiflora were identified.The PlCDPK24 gene was cloned from its stigma and bioinformatics analysis was performed on the gene structure and physicochemical properties of the encoded proteins.The intracellular local-ization of PlCDPK24 was determined,and its expression pattern during pollen tube growth in the stigma was assessed using RT-qPCR.Additionally,its response to Ca2+and the Ca2+inhibitor ethylene glycol tetraacetic acid(EGTA)was analyzed.[Result]Fifteen PlCDPK genes that were highly expressed in hybrid unaffinity combinations were identified and screened,and the coding regions of the genes all contained STKc-CAMK structural domains,EF-hand chiral structural domains,and motifs 1 to 10.Among them,the sequence of the Paeonia lactiflora PlCDPK24 gene,which was homologous to the pollen-tube-growth-associated AtCDPK24 that had been reported in Arabidopsis thaliana,was complete,with an open reading frame of 1 599 bp and encoding a 5.69 theoretical iso-electric point,and 532 amino acids,characterized by a negatively charged hydrophilic protein.The PlCDPK24 protein is localised in the nucleus and has the highest homology with the peony PsCDPK24 protein.The results of real-time fluorescence quantification showed that PlCDPK24 was highly expressed in peony stigmas compared with other parts of peony;The expression of PlCDPK24 was sig-nificantly up-regulated in the late hybridisation stage(24 and 36 h)compared with the self-cross combination,which was significantly higher than that in the late self-crossing stage;The expression of PlCDPK24 was significantly up-regulated in the hybrid stigmas treated with Ca2+as well as with the Ca2+inhibitor,EGTA,relative to the The expression of PlCDPK24 was significantly up-regulated in self-crossed stigmas,and both Ca2+and EGTA treatments reduced the expression of PlCDPK24 to dif-ferent degrees.[Conclusion]PlCDPK24 was highly expressed in the stigma of Paeonia lactis,and increased with time after cross-pollination.The expression level of PlCDPK24 was significantly lower than that of the self-pollinated stigma,suggesting that PlCDPK24 was involved in regulating the growth and elongation of pollen tubes in the stigma to a large extent.关键词
芍药/远缘杂交不亲和/钙依赖性蛋白激酶/基因克隆/花粉生长Key words
Paeonia lactiflora/distant hybridization incompatibility/calcium-dependent protein kinase/gene cloning/pollen growth分类
农业科技引用本文复制引用
贺丹,尤啸龙,周平西,华超,郭浩楠,张明星,王警琨,何松林..芍药钙依赖性蛋白激酶基因PlCDPK24的鉴定及表达分析[J].河南农业大学学报,2025,59(4):602-613,12.基金项目
国家自然科学基金项目(32371952) （32371952）
河南省自然科学基金项目(232300420006) （232300420006）
河南省科技攻关项目(242102110256) （242102110256）
河南省高等学校重点科研项目(23A220001) （23A220001）
