香蕉Ca^(2+)-ATPase基因家族的鉴定及功能分析  

Identification and Functional Analysis of Ca^(2+)-ATPase Gene Family in Banana

作　　者：滕梦鑫 徐亚 何静 汪奇 乔飞[2] 李敬阳 李新国 TENG MengXin;XU Ya;HE Jing;WANG Qi;QIAO Fei;LI JingYang;LI XinGuo(School of Tropical Agriculture and Forestry,Hainan University/National Key Laboratory of Tropical Crop Biological Breeding,Haikou 570228;Chinese Academy of Tropical Agricultural Sciences Tropical Crops Genetic Resources Institute,Haikou 571101)

机构地区：[1]海南大学热带农林学院/热带作物生物育种全国重点实验室,海口570228 [2]中国热带农业科学院热带作物品种资源研究所,海口571101

出　　处：《中国农业科学》2025年第7期1418-1433,I0001,I0002,共18页Scientia Agricultura Sinica

基　　金：国家自然科学基金(32160679,31760549)。

摘　　要：【目的】Ca^(2+)-ATPase是植物中重要的Ca^(2+)外排通道,对于维持细胞稳态、诱导植物对胁迫的响应等方面起着重要的作用。利用生物信息学和分子生物学等手段,对香蕉A基因组中Ca^(2+)-ATPase家族成员进行分析,检测基因表达量的变化,并验证基因功能,为探究香蕉耐盐机制提供参考。【方法】通过基因家族名称并结合拟南芥和水稻Ca^(2+)-ATPase序列,在香蕉基因组数据库中BLAST香蕉A基因组中该家族成员;利用Pfam、ExPASy、Cell-PLoc、NCBI、MEGA-X、TBtools等在线网站或软件,预测分析香蕉Ca^(2+)-ATPase的蛋白理化性质、亚细胞定位、保守结构域、顺式作用元件等;采用qRT-PCR技术该家族基因的表达情况,对关键基因进行亚细胞定位验证和原核表达分析。【结果】在香蕉A基因组鉴定到20个Ca^(2+)-ATPase家族成员,其中,有13个MaACAs和7个MaECAs;蛋白理化性质分析显示,Ca^(2+)-ATPase含有6—11个不等的跨膜结构,编码857—1 103个氨基酸,亚细胞定位预测MaACAs可能定位在细胞质膜、叶绿体、内质网及液泡膜上,MaECAs均定位于内质网膜;保守基序高度一致,除MaACA13外均含有该家族特有的4个保守结构域;含有光、激素、防御和应激等响应元件。qRT-PCR结果表明,100 mmol·L^(-1) NaCl处理巴西蕉细胞,除MaECA3外,Ca^(2+)-ATPase基因家族其他成员在处理期间均出现了不同程度的上调表达,其中MaACA5和MaACA10的上调幅度较大,并且这两个基因的表达均受Ca^(2+)影响。烟草叶片亚细胞定位结果显示,MaACA5和MaACA10定位在细胞质膜上;重组质粒pET28a-MaACA5、pET28a-MaACA10和转入大肠杆菌BL21中,诱导表达后的重组菌株在800 mmol·L^(-1) NaCl、800mmol·L^(-1)甘露醇和50℃高温条件下的生长情况优于对照菌株。【结论】在香蕉A基因组中鉴定到20个Ca^(2+)-ATPase家族成员,基因结构具有较高的保守性,分布在10条染色体上,包含激素、防御和应激相关的�【Objective】Ca^(2+)-ATPase is a vital Ca^(2+)efflux channel in plants,which plays an important role in maintaining cell homeostasis and inducing plant response to stress.In this study,bioinformatics and molecular biology were used to analyze the members of Ca^(2+)-ATPase family in banana A genome,detect the changes of gene expression and verify gene function,so as to provide reference for exploring the mechanism of banana salt tolerance.【Method】BLAST the members of banana A genome in banana genome database by gene family name combined with Arabidopsis and rice Ca^(2+)-ATPase sequences,and use Pfam,ExPASy,Cell-PLoc,NCBI,MEGA-X,TBtools and other online websites or software to predict and analyze the protein physicochemical properties,subcellular localization,conservative domain and cis-acting elements of banana Ca^(2+)-ATPase.The expression of the family genes was analyzed by qRT-PCR,and the subcellular localization and prokaryotic expression of the key genes were analyzed.【Result】In this study,20 members of Ca^(2+)-ATPase family were identified in banana A genome,including 13 MaACAs and 7 MaECAs;proteins analysis of physicochemical properties showed that Ca^(2+)-ATPase contained 6-11 transmembrane structures,encoding amino acids between 857 and 1103.Subcellular localization predicted that MaACAs might be located on plasma membrane,chloroplast,endoplasmic reticulum and tonoplast,while MaECAs was located in endoplasmic reticulum.The conserved motifs are highly consistent,including four conserved domains peculiar to the family except MaACA13,responses to light,hormone,defense and stress.qRT-PCR results showed that other members of Ca^(2+)-ATPase gene family except MaECA3 were up-regulated in Brazilian banana cells treated with 100 mmol·L^(-1) NaCl,in which MaACA5 and MaACA10 were up-regulated to a large extent,and the expression of these two genes was affected by Ca^(2+).The results of subcellular localization of tobacco leaves showed that MaACA5 and MaACA10 were located on the plasma membrane,and the rec

关 键 词：巴西蕉 盐胁迫 钙离子 Ca^(2+)-ATPase 表达分析 基因克隆 亚细胞定位 

分 类 号：S668.1[农业科学—果树学]