[1]姜玉素,李珍,王庆莲,等.草莓KEA家族基因的克隆、鉴定及表达分析[J].江苏农业学报,2019,(02):391-399.[doi:doi:10.3969/j.issn.1000-4440.2019.02.021]
　JIANG Yu-su,LI Zhen,WANG Qing-lian,et al.Cloning, characterization and expression analysis of KEA family genes in strawberry[J].,2019,(02):391-399.[doi:doi:10.3969/j.issn.1000-4440.2019.02.021]
点击复制

草莓KEA家族基因的克隆、鉴定及表达分析()
分享到：

江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2019年02期
页码:
391-399
栏目:
园艺
出版日期:
2019-04-30

文章信息/Info

Title:
Cloning, characterization and expression analysis of KEA family genes in strawberry
作者:
姜玉素1; 2; 李珍2; 王庆莲3; 赵密珍3; 宋志忠2; 3
（1.烟台职业学院，山东烟台264670；2.鲁东大学农学院，山东烟台264025；3.江苏省农业科学院果树研究所，江苏南京210014）
Author(s):
JIANG Yu-su1; 2; LI Zhen2; WANG Qing-lian3; ZHAO Mi-zhen3; SONG Zhi-zhong2; 3
(1.Yantai Vocational College, Yantai 264670, China；2.School of Agriculture, Ludong University, Yantai 264025, China；3.Institute of Pomology, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China)
关键词:
草莓; K+/H+逆向转运体; 生物信息学分析; 基因克隆与表达
Keywords:
strawberry; K+/H+ antiporter; bioinformatics analysis; gene cloning and expression
分类号:
S668.4
DOI:
doi:10.3969/j.issn.1000-4440.2019.02.021
文献标志码:
A
摘要:
K+/H+逆向转运体（KEA）介导细胞中K+和H+的动态平衡，在维持植物体内的离子平衡、生长发育和信号转导中起重要作用，然而，相关研究主要体现在模式作物拟南芥中，果树中KEA家族基因的功能依然未知。本研究以Yellow Wonder 5AF8草莓为材料，筛选并克隆KEA家族基因，并对其进行生物信息学鉴定和表达特征分析，为研究果树K+/H+平衡及钾素动态平衡机制提供基因资源和理论依据。结果表明,在草莓基因组中检索并克隆到5个KEA家族基因，命名为FveKEA1～FveKEA5，属于典型的植物K+/H+逆向转运体基因；编码的蛋白质与7种已报道的不同科属植物的KEA家族蛋白在氨基酸水平上具有25.00%的一致性，并可分为2个亚族（Group I和Group II），其中，草莓FveKEA1和FveKEA2属于Group I，只含有4个Motif基序，而FveKEA3～FveKEA5属于Group II，含有7个Motif基序；系统进化树表明草莓FveKEA2和FveKEA4分别与葡萄VvKEA2和苹果MdoKEA6紧密聚在一起，草莓FveKEA1、FveKEA3和FveKEA5分别与葡萄VvKEA1、白杨PtrKEA4、桃PpeKEA4等相应成员在遗传距离上较近；草莓KEA家族蛋白主要定位于细胞质膜，均含有12～14个跨膜区，除FveKEA3外，均为稳定蛋白，且只有FveKEA5含有信号肽；转录表达谱分析结果揭示草莓KEA家族基因在多种组织或器官中均有表达，实时荧光定量PCR结果表明FveKEA1在5AF8草莓不同组织中的整体表达水平最高，在花瓣和未成熟果实中的表达量最为突出，其次是FveKEA4，而其他3个基因的整体表达水平相对较低。此外，在草莓KEA基因启动子区域鉴定到至少16种顺式作用元件，且均含有光感应、胚乳表达和脱落酸（ABA）响应的作用元件。
Abstract:
In plants, K+ efflux antiporters (KEAs) mediate the cellular K+/H+ homeostasis and play key role in the ion balance, growth and development and signal transduction. However, the relevant studies mainly focused on model plant Arabidopsis, the functions of KEA family genes in fruit trees were rarely known. Using Yellow Wonder 5AF8 strawberry as material, KEA family genes bioinformatics identification and gene expression characteristics were analyzed to provide gene resources and theoretical foundation for the study of K+/H+ balance and K+ homeostasis mechanisms in fruit trees. Results showed that five KEA family genes were isolated from strawberry, named FveKEA1-FveKEA5, which belonged to the typical plant K+/H+ antiporter gene. The encoded proteins shared an overall identity of 25.00% at the amino acid level with the KEA family proteins of seven different families, and could be divided into two subgroups, Group I and Group II. FveKEA1 and FveKEA2 belonged to Group I, and contained four Motif motifs. FveKEA3-FveKEA5 belonged to Group II, and contained seven Motif motifs. Phylogenetic tree analysis showed that FveKEA2 and FveKEA4 of strawberry were closely related to VvKEA2 of grape and MdoKEA6 of apple, respectively, and FveKEA1, FveKEA3 and FveKEA5 were closely related to VvKEA1 of grape, PtrKEA4 of polar and PpeKEA4 of peach, correspondingly. All FveKEA proteins were majorly localized in plasma membrane and contained 12-14 transmembrane domains (TMs). All strawberry KEA proteins were stable protein with the exception of FveKEA3, and only FveKEA5 possessed the signal peptide. The results of transcriptional profiling revealed that FveKEA genes could be detected in various tissues or organs of strawberry. qRT-PCR analysis results indicated that FveKEA1 had the highest expression level in different tissues of 5AF8 strawberry on the whole and the most prominent expression level in petals and immature fruits, followed by FveKEA4, while the overall expression levels of the other three genes were relatively low. Moreover, at least 16 cis-acting elements were identified in the promoter region of strawberry KEA family genes, and all of them contained elements of light sensing, endosperm expression and abscisic acid response.

参考文献/References:

[1]VRY A A, SENTENAC H. Molecular mechanisms and regulation of K+ transport in higher plants [J]. Annual Review of Plant Biology, 2003, 54: 575-603.
[2]GRABOV A. Plant KT/KUP/HAK potassium transporters: Single family-multiple functions [J]. Annals of Botany, 2007, 99: 1035-1041.
[3]DEMIRAL M A, KSEOGLU A T. Effect of potassium on yield, fruit quality, and chemical composition of green house-grown aalia melon [J]. Journal of Plant Nutrition, 2005, 28: 93-100.
[4]YURTSEVEN E, KESMEZ G D, NLKARA A. The effects of water salinity and potassium levels on yield, fruit quality and water consumption of a native central anatolian tomato species (Lycopersicon esculantum) [J]. Agricultural Water Management, 2005, 78: 128-135.
[5]HARTZ T K, JOHNSTONE P R, FRANCIS D M, et al. Processing tomato yield and fruit quality improved with potassium fertigation [J]. Hort Science, 2005, 40: 1862-1867.
[6]宋志忠,郭绍雷,马瑞娟,等. KT/HAK/KUP家族基因在桃开花期的表达及对钾肥施放的响应分析 J]. 中国农业科学, 2015, 48(6): 1177-1185.
[7]宋志忠,许建兰,张斌斌,等. 叶面喷施钾肥对霞脆桃果实品质及KUP基因表达的影响[J]. 江苏农业学报，2018，34（5）：1107-1112.
[8]CHANROJ S, LU Y, PADMANABAN S, et al. Plant-specific cation/H+ exchanger 17 and its homologs are endomembrane K+ transporters with roles in protein sorting [J]. Journal of Biology Chemistry, 2011, 286(39): 33931-33941.
[9]CHANROJ S, WANG G, VENEMA K, et al. Conserved and diversified gene families of monovalent cation/H+ antiporters from algae to flowering plants [J]. Frontiers in Plant Science, 2012, 3: 25.
[10]BASSIL E, BLUMWALD E. The ins and outs of intracellular ion homeostasis: NHX-type cation/H+ transporters [J]. Current Opinion in Plant Biology, 2014, 22: 1-6.
[11]MASER P, THOMINE S, SCHROEDER J I, et al. Phylogenetic relationships within cation transporter families of Arabidopsis [J]. Plant Physiology, 2001, 126: 1646-1667.
[12]HAN L, LI J L, WANG L, et al. Identification and localized expression of putative K+/H+ antiporter genes in Arabidopsis [J]. Acta Physiologiae Plantarum, 2015, 37(5): 1-14.
[13]ARANDA-SICILIA M N, CAGNAC O, CHANROJ S, et al. Arabidopsis KEA2, a homolog of bacterial KefC, encodes a K+/H+ antiporter with a chloroplast transit peptide [J]. BBA-Biomembranes, 2012, 1818(9): 2362-2371.
[14]ZHENG S, PAN T, FAN L G, et al. A novel AtKEA gene family, homolog of bacterial K+/H+ antiporters, plays potential roles in K+ homeostasis and osmotic adjustment in Arabidopsis [J]. PLoS ONE, 2013, 8(11): e81463.
[15]ARMBRUSTER U, CARRILLO L R, VENEMA K, et al. Ion antiport accelerates photosynthetic acclimation in fluctuating light environments [J]. Nature Communications, 2014, 5: 5439.
[16]KUNZ H H, GIERTH M, HERDEAN A, et al. Plastidial transporters KEA1, -2, and -3 are essential for chloroplast osmoregulation, integrity, and pH regulation in Arabidopsis [J]. Proceedings of the National Academy of Sciences of the United States of America, 2014, 111(20): 7480-7485.
[17]ZHOU H, QI K, LIU X, et al. Genome-wide identification and comparative analysis of the cation proton antiporters family in pear and four other Rosaceae species [J]. Molecular Genetics and Genomics, 2016, 291: 1727-1742.
[18]KANG C, DARWISH O, GERETZ, et al. Genome-scale transcriptomic insights into early-stage fruit development in woodland strawberry Fragaria vesca [J]. Plant Cell, 2013, 25:1960-1978.
[19]FAIT A, HANHINEVA K, BELEGGIA R, et al. Reconfiguration of the achene and receptacle metabolic networks during strawberry fruit development [J]. Plant Physiology, 2008, 148 (2):730-750.
[20]LIVAK K J, SCHMITTGEN T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2-△△Ct method [J]. Methods, 2001, 25(4): 402-408.
[21]WANG Y, WU W H. Genetic approaches for improvement of the crop potassium acquisition and utilization efficiency [J]. Current Opinion in Plant Biology, 2015, 25:46-52.
[22]CHEN H T, CHEN X, WU B Y, et al. Whole-genome identification and expression analysis of K+ efflux antiporter (KEA) and Na+/H+ antiporter (NHX) families under abiotic stress in soybean [J]. Journal of Intergrative Agriculture, 2015, 14(6): 1171-1183.
[23]SONG Z Z, GUO S L, ZHANG C H, et al. KT/HAK/KUP potassium transporter genes differentially expressed during fruit devcelopment, ripening, and postharvest shelf-life of ‘Xiahui6’ peaches [J]. Acta Physiologiae Plantarum, 2015, 37:131.

相似文献/References:

[1]巫建华,贾思振,冯英娜,等.茉莉酸合成关键酶基因FaOPR3调控草莓果实成熟[J].江苏农业学报,2016,(05):1148.[doi:10.3969/j.issn.1000-4440.2016.05.031]
　WU Jian-hua,JIA Si-zhen,FENG Ying-na,et al.Regulation of strawberry fruit ripening by jasmonic acid synthesis gene FaOPR3[J].,2016,(02):1148.[doi:10.3969/j.issn.1000-4440.2016.05.031]
[2]赵密珍,庞夫花,袁华招,等.不同栽培条件下草莓品种宁玉花序分化进程[J].江苏农业学报,2016,(01):196.[doi:10.3969/j.issn.1000-4440.2016.01.030 ]
　ZHAO Mi-zhen,PANG Fu-hua,YUAN Hua-zhao,et al.Floral bud differentiation of strawberry Ningyu under conditions of forcing cultivation and open field cultivation[J].,2016,(02):196.[doi:10.3969/j.issn.1000-4440.2016.01.030 ]
[3]吴祥,姚克兵,吉沐祥,等.句容地区草莓枯萎病病原菌的分离鉴定及田间防治[J].江苏农业学报,2015,(04):764.[doi:10.3969/j.issn.1000-4440.2015.04.009]
　WU Xiang,YAO Ke-bing,JI Mu-xiang,et al.Isolation and identification and field control of strawberry fusarium wilt in Jurong, Jiangsu province[J].,2015,(02):764.[doi:10.3969/j.issn.1000-4440.2015.04.009]
[4]周游,李海梅,赵金山,等.乳酸菌对草莓生长和品质性状的影响[J].江苏农业学报,2017,(05):1124.[doi:doi:10.3969/j.issn.1000-4440.2017.05.025]
　ZHOU You,LI Hai-mei,ZHAO Jin-shan,et al.Effects of lactic acid bacteria on the growth and quality characters of strawberry[J].,2017,(02):1124.[doi:doi:10.3969/j.issn.1000-4440.2017.05.025]
[5]江景勇,王会福,何玲玲,等.台州草莓农药残留风险评估[J].江苏农业学报,2017,(06):1408.[doi:doi:10.3969/j.issn.1000-4440.2017.06.030]
　JIANG Jing-yong,WANG Hui-fu,HE Ling-ling,et al.Risk assessment of pesticide residues in strawberry from Taizhou city[J].,2017,(02):1408.[doi:doi:10.3969/j.issn.1000-4440.2017.06.030]
[6]丛嘉昕,宋江峰,李大婧,等.基于RSM和MLP-ANN的草莓果浆超声酶解参数优化[J].江苏农业学报,2018,(06):1354.[doi:doi:10.3969/j.issn.1000-4440.2018.06.022]
　CONG Jia-xin,SONG Jiang-feng,LI Da-jing,et al.Optimization of ultrasonic enzymolysis of strawberry pulp based on RSM and MLP-ANN[J].,2018,(02):1354.[doi:doi:10.3969/j.issn.1000-4440.2018.06.022]
[7]王廷峰,赵密珍,关玲,等.玉米套作及秸秆还田对草莓连作土壤养分及微生物区系的影响[J].江苏农业学报,2019,(06):1421.[doi:doi:10.3969/j.issn.1000-4440.2019.06.022]
　WANG Ting-feng,ZHAO Mi-zhen,GUAN-Ling,et al.Effects of intercropping with corn and straw returning on nutrients and microflora in strawberry continuous cropping soil[J].,2019,(02):1421.[doi:doi:10.3969/j.issn.1000-4440.2019.06.022]
[8]张宽朝,陈杰,马伟,等.响应面法优化草莓总黄酮提取工艺[J].江苏农业学报,2019,(06):1450.[doi:doi:10.3969/j.issn.1000-4440.2019.06.026]
　ZHANG Kuan-chao,CHEN Jie,MA Wei,et al.Optimization on extraction technology of total flavonoids from strawberry by response surface methodology[J].,2019,(02):1450.[doi:doi:10.3969/j.issn.1000-4440.2019.06.026]
[9]李梦雅,陈莎莎,王世梅.连作草莓低发病土壤与高发病土壤理化性质及生物学特征差异比较[J].江苏农业学报,2021,(04):910.[doi:doi:10.3969/j.issn.1000-4440.2021.04.013]
　LI Meng-ya,CHEN Sha-sha,WANG Shi-mei.Contrast of physicochemical properties and biological characteristics of low disease and high disease soils of continuous cropping strawberry[J].,2021,(02):910.[doi:doi:10.3969/j.issn.1000-4440.2021.04.013]
[10]朱丽,王庆莲,唐山远,等.不同试剂处理对草莓植株生长及根际微生物群落结构的影响[J].江苏农业学报,2023,(01):198.[doi:doi:10.3969/j.issn.1000-4440.2023.01.023]
　ZHU Li,WANG Qing-lian,TANG Shan-yuan,et al.Effects of different reagent treatments on strawberry plant growth and rhizosphere microbial community structure[J].,2023,(02):198.[doi:doi:10.3969/j.issn.1000-4440.2023.01.023]

备注/Memo

备注/Memo:
收稿日期：2018-07-16 基金项目：国家自然科学基金项目（31501743、31301764）；江苏省农业重大新品种创制项目（PZCZ201721）；泰州市科技支撑计划项目（TN201708） 作者简介：姜玉素（1973-），女，山东烟台人，硕士，讲师，主要进行食品发酵与分子生物学研究。（Tel）13863857817;（E-mail）jiangyusu906552@163.com 通讯作者：宋志忠， （Tel）025-84390255；（E-mail）szhzh2000@163.com
更新日期/Last Update: 2019-05-05