[1]金 勋,杨 柳,潘红丽,等.大豆GmFtsH2基因的克隆及表达分析[J].黑龙江农业科学,2022,(01):6-19.[doi:DOI:10.11942/j.issn1002-2767.2022.01.0006]
　JIN Xun,YANG Liu,PAN Hong-li,et al.Cloning and Expression Analysis of GmFtsH2 Genes in Soybean[J].HEILONGJIANG AGRICULTURAL SCIENCES,2022,(01):6-19.[doi:DOI:10.11942/j.issn1002-2767.2022.01.0006]
点击复制

大豆GmFtsH2基因的克隆及表达分析

《黑龙江农业科学》[ISSN:1002-2767/CN:23-1204/S] 卷: 期数: 2022年01 页码: 6-19 栏目: 出版日期: 2022-01-10
Title:
Cloning and Expression Analysis of GmFtsH2 Genes in Soybean
作者:
金 勋; 杨 柳; 潘红丽; 芮海英; 王丽娜; 李 娜; 李泽宇
黑龙江省农业科学院 大庆分院，黑龙江 大庆 163316
Author(s):
JIN Xun; YANG Liu; PAN Hong-li; RUI Hai-ying; WANG Li-na; LI Na; LI Ze-yu
Daqing Branch， Heilongjiang Academy of Agricultural Sciences,Daqing 163316，China
关键词:
大豆; FtsH; 逆境胁迫; 表达分析
Keywords:
soybean;  FtsH;  abiotic stress;  expression analysis
DOI:
DOI:10.11942/j.issn1002-2767.2022.01.0006
文献标志码:
A
摘要:
为了进一步了解FtsH基因在大豆中的表达特性和功能，通过GenBank中拟南芥等其他植物FtsH基因序列设计1对简并引物，从盐处理的大豆叶片中克隆GmFtsH2的全长cDNA序列，并利用在线生物信息学软件对其进行分析。采用实时荧光定量PCR方法分析GmFtsH2在低温、盐、干旱和高温处理下的表达情况。从大豆中克隆到一条全长为2 200 bp的mRNA序列，命名为GmFtsH2基因，开放阅读框2 091 bp，编码696个氨基酸，相对分子量约为75.089 kDa，理论等电点为5.65，蛋白质不稳定系数为34.90，是稳定蛋白。大豆GmFtsH2与来源于三叶草、葡萄、水稻、玉米和拟南芥类的FtsH类同源蛋白的氨基酸序列比对发现，这些蛋白具有FtsH蛋白共同的保守结构域，包括N端两个跨膜域、AAA结构域、锌离子结合模块等。大豆GmFtsH2与其他19种植物FtsH 蛋白的系统进化分析表明，与野生豆和菜豆两个蛋白的亲缘关系较近。RT-PCR分析表明，GmFtsH2在大豆的叶片、茎、根、茎尖的组织内均有表达，且在茎尖中表达量最高，茎中最低。大豆叶片在4 ℃低温和NaCl处理下，GmFtsH2的表达随时间延长逐渐被诱导，分别在处理后的9 h和12 h达到最高峰；而经干旱诱导的大豆叶片，GmFtsH2的表达在第3天达到最高峰，随后GmFtsH2的表达下调，第11天又达到一个小高峰。推测GmFtsH2在大豆抵御逆境胁迫中起作用。
Abstract:
In order to further understand the expression characteristics and functions of FtsH gene in soybean,primers for cloning GmFtsH2 gene were designed in references with the FtsH gene sequences of Arabidopsis and other plants opened in GenBank.Full-length cDNA of GmFtsH2 was cloned from soybean leaves with salt treatment,and analyzed with bioinformatics software online.The expression of GmFtsH2 under the low temperature,salinity,drought and high temperature conditions were analyzed with real-time PCR.A full-length mRNA of 2 200 bp was cloned from soybean,and named GmFtsH2 gene.The open reading frame was 2 091 bp,encoding 696 amino acids.The relative molecular weight was about 75.089 kDa and the theoretical isoelectric point was 5.65.Its protein instability index was 34.90,indicting a stable protein.The alignment of GmFtsH2 in soybean and homologous protein amino acid sequence of FtsH from clover,grape,rice,corn and Arabidopsis sequence revealed that those proteins showed a common conservative domain structure,including membrane-spanning domains to N terminus,AAA-structure domain,Zinc ions in combination with module,etc.Phylogenetic analysis between GmFtsH2 in soybean and FtsH protein of the other 19 kinds of plants showed that it was resembled to wild beans and kidney beans.Real-time RT-PCR analysis showed that GmFtsH2 was expressed in soybean leaves,stem,root,stem tip,and the stem tip had the highest expression,the lowest expression was in the stem.The expression of GmFtsH2 in soybean leaves was induced gradually with time and to a maximum at 9 and 12 h respectively under 4 ℃ low temperature and NaCl treatment.Under drought treatment GmFtsH2 expression reached to maximum after 3 days.Thereafter GmFtsH2 were down expressed subsequently and reached to a small peak again after 11 days treatment.It is suggested that the GmFtsH2 plays a role in the resistance of adversity stress in soybean.

参考文献/References:

参考文献：

［1］KARATA K,INAGAWA T,WILKINSON A J,et al.Dissecting the role of a conserved motif(the second region of homology)in the AAA family of ATPases.Site-directed mutagenesis of the ATP-dependent protease FtsH［J］.Journal of Biologlcal Chemistry,1999,274:26225-26232.

［2］LANGER T.AAA proteases:Cellular machines for degrading membrane proteins［J］.Trends in Biochemical Sciences,2000,25:247-251.

［3］OGURA T,WILKINSON A J.AAA+superfamily ATPases:Common structure-diverse function［J］.Genes Cells,2001,6:575-597.

［4］MIHO Y N,YASUSI Y.Quality control of Photosystem II:The molecular basis for the action of FtsH protease and the dynamics of the thylakoid membranes［J］.Journal of Photochemistry and Photobiology B:Biology,2014 (137):100-106.

［5］NEUWALD A F,ARAVIND L,SPOUGE J L,et al.AAA+:A class of chaperone-like ATPases associated with the assembly,operation,and disassembly of protein complexes［J］.Genome Research,1999,9(1):27-43. 

［6］SUMMER E J,CLINE K.The nucleotide sequence of a tobacco homolog of Pftf(accession no.AF117339):Evidence for two plastid-localized AAA-family proteins in higher plants(PGR 99-029)［J］.Plant Physiology,1999,119(3):1147-1147.

［7］SAKAMOTO W,ZALTSMAN A,ADAM Z,et al.Coordinated regulation and complex formation of YELLOW VARIEGATED1 and YELLOW VARIEGATED2,chloroplastic FtsH metalloproteases involved in the repair cycle of photosystem II in Arabidopsis thylakoid membranes［J］.Plant Cell,2003,15:2843-2855. 

［8］ADAM Z,ADAMSKA I,NAKABAYASHI K,et al.Chloroplast and mitochondrial proteases in Arabidopsis.A proposed nomenclature［J］.Plant Physiology,2001,125:1912-1918. 

［9］LINDAHL M,TABAK S,CSEKE L,et al.Identification,characterization,and molecular cloning of a homologue of the bacterial FtsH protease in chloroplasts of higher plants［J］.Journal of Biologlcal Chemistry,1996,271:29329-29334.

［10］DEUERLING E,PAESLACK B,SCHUMANN W.The FtsH gene of Bacillus subtilis is transiently induced after smotic and temperature upshift［J］.Journal of Bacteriology,1995,177:4105-4112.

［11］DEUERLING E,MOGK A,RICHTER C,et al.The FtsH gene of Bacillus subtilis is involved in major cellular processes such as sporulation,stress adaptation and secretion［J］.Molecular Microbiology,1997,23:921-933.

［12］IVASHUTA S,IMAI R,UCHIYAMA K,et al.Changes in chloroplast FtsH-like gene during cold acclimation in alfalfa(Medicago sativa)［J］.Journal of Plant Physiology,2002,159:85-90. 

［13］BOURDINEAUD J P,NEHME B,TESSE S,et al.The FtsH gene of the wine bacterium Oenococcus oeni is involvedin protection against environmental stress［J］.Applied and Environmental Microbiology,2003，69:2512-2520.

［14］CHEN M,CHOI Y,VOYTAS D F,et al.Mutations in the Arabidopsis VAR2 locus cause leaf variegation due to the loss of a chloroplast FtsH protease［J］.The Plant Journal,2000,22:303-313. 

［15］DUWAT P,EHRLICH S D,GRUSS A.The recA gene of Lactococcus lactis:Characterization and involvement in oxidative and thermal stress［J］.Molecular Microbiology,1995,17:1121-1131.

［16］GUO Z H,GAO X,CAI H Q,et al.Genome-wide identification,evolution and expression analysis of the FtsH gene during fruit development in pear(Pyrus bretschneideri)［J］.Plant Biotechnology Reports,2021,15:537-550.

［17］SAKAMOTO W,ZALTSMAN A,ADAM Z,et al.Coordinated regulation and complex formation of YELLOW VARIEGATED1 and YELLOW VARIEGATED2,chloroplastic FtsH metalloproteases involved in the repair cycle of photosystem II in Arabidopsis thylakoid membranes［J］.Plant Cell,2003,15:2843-2855.

［18］LINDAHL M,SPETEA C,HUNDAL T,et al.The thylakoid FtsH protease plays arole in the light-induced turnover of the photosystem II D1 protein［J］.Plant Cell,2000,12:419-431.

［19］YOSHIOKA M,UCHIDA S,MORI H,et al.Quality control of photosystem II［J］.Journal of Biologlcal Chemistry,2006,281:21660-21669.

［20］LIU X Y,RODERMEL S R,YU F.Avar2 leaf variegation suppressor locus suppressor of variegation3,encodes a putative chloroplast translation elongation factor that is important for chloroplast development in the cold［J］.BMC Plant Biology,2010,10:1-18. 

［21］SILVA P,THOMPSON E,BAILEY S,et al.FtsH is involved in the early stages of repair of photosystem II in synechocystis sp PCC 6803［J］.Plant Cell,2003,15(9):2152-2164.

［22］SINGH M,YAMAMOTO Y,SATOHA K,et al.Post-illumination-related loss of photochemical efficiency of Photosystem II and degradation of the D1 protein are temperature-dependent［J］.Journal of Plant Physiology,2005,162(11):1246-1253. 

［23］TIAN Y N,ZHONG R H,WEI J B,et al.Arabidopsis chlorophyllase 1 protects young leaves from long-term photodamage by facilitating FtsH-mediated D1 degradation in photosystem II repair［J］.Molecular Plant,2021,14(7):1149-1167.

［24］CHEN J,BURKE J J,VELTEN J,et al.FtsH11 protease plays a critical role in Arabidopsis thermotolerance［J］.Journal Plant,2006,48(1):73-84.

［25］SEO S,OKAMOTO M,IWAI T,et al.Reduced levels of chloroplast FtsH protein in tobacco mosaic virus-infected tobacco leaves accelerate the hypersensitive reaction［J］.Plant Cell,2000,12:917-932.

［26］IVASHUTA S,IMAI R,UCHIYAMA K,et al.Changes in chloroplast FtsH-like gene during cold acclimation in alfalfa(Medicago sativa)［J］.Journal Plant Physiology,2002,159:85-90.

［27］SUN A Q,YI S Y,YANG J Y,et al.Identification and characterization of a heat-inducible FtsH gene from tomato(Lycopersicon esculentum Mill.)［J］.Plant Science,2006,170:551-562.

［28］范敏,金黎平,黄三文,等.马铃薯SoFtsH基因全长cDNA克隆与在干旱条件下表达研究［J］.作物学报,2007(11):1748-1754. 

［29］HANNA J,MALGORZATA K,JOANNA S.Protein quality control in organelles-AAA/FtsH story［J］.Biochimica et Biophysica Acta-Molecular Cell Research,2013,1833:381-387. 

［30］FISCHER B,RUMMEL G,ALDRIDGE P,et al.The FtsH protease is involved in development,stress response and heat shock control in Caulobacter crescentus［J］.Molecular Microbiology,2002,44:461-478. 

［31］YIN Z T,MENG F F,SONG H N,et al.GmFtsH9 expression correlates with in vivo photosystem II function:Chlorophyll a fluorescence transient analysis and eQTL mapping in soybean［J］.Planta,2011,234(4):815-827.

［32］李永春,张春艳,张宁,等.小麦质膜蛋白基因TaPM19-1的克隆及其对非生物胁迫的响应［J］.中国农业科学,2012,45(12):2502-2509. 

［33］SAKAMOTO W,TAMURA T,HANBA-TOMITA Y,et al.The VAR1 locus of Arabidopsis encodes a chloroplastic FtsH and is responsible for leaf variegation in the mutant alleles［J］.Genes Cells,2002,7:769-780.

［34］LIU X Y,YU F,RODERMEL S.Arabidopsis chloroplast FtsH,var2 and suppressor of var2 leaf variegation:A review［J］.Journal of Integrative Plant Biology,2010,52:750-761.

［35］YUE G D,HU X R,HE Y,et al.Identification and characterization of two members of the FtsH gene family in maize(Zea mays L.)［J］.Molecular Biology Reports,2010,37:855-863.

［36］李亮,李泽宇,王丽娜,等.不同大豆品种耐盐性表现分析［J］.黑龙江农业科学,2013(2):17-20.

［37］芮海英,王丽娜,金铃,等.苗期干旱胁迫对不同大豆品种叶片保护酶活性及丙二醛含量的影响［J］.大豆科学,2013，32(5):647-649.

相似文献/References:

[1]吕静,韩莉梅,张所有.超氧化物歧化酶模拟物（SODM）对大豆保护酶系统的影响[J].黑龙江农业科学,2013,(01):23.
　LYUJing,HANLi-mei,ZHANGSuo-you.EffectsofSuperoxideDismutaseMimics(SODM)onSoybeanProtectiveEnzymeSystem[J].HEILONGJIANG AGRICULTURAL SCIENCES,2013,(01):23.
[2]吴俊彦,于晓光.黑河地区灾后补种大豆品种筛选试验[J].黑龙江农业科学,2014,(07):13.
　WU Jun yan,YU Xiao guang.Soybean Varieties Screening Test for Reseeding in Heihe Region after Disaster[J].HEILONGJIANG AGRICULTURAL SCIENCES,2014,(01):13.
[3]安咏梅,王家军,李进荣,等.大豆抗胞囊线虫的分子标记研究[J].黑龙江农业科学,2014,(07):15.
　AN Yong mei,WANG Jia jun,LI Jin rong,et al.Molecular Marker Assisted Selection on Soybean Resistant to Soybean Cyst Nematode[J].HEILONGJIANG AGRICULTURAL SCIENCES,2014,(01):15.
[4]刘兴龙,王克勤,朱立新,等.大豆蚜对大豆阶段性危害研究[J].黑龙江农业科学,2014,(06):57.
　LIU Xinglong,WANG Keqin,ZHU Lixin,et al.Study on Effect of Soybean Aphid on Soybean Growth at Different Stages[J].HEILONGJIANG AGRICULTURAL SCIENCES,2014,(01):57.
[5]赵 寅.富裕县大豆生产现状及发展建议[J].黑龙江农业科学,2014,(05):150.
　ZHAO Yin.Production Situation and Suggestions of Soybean in Fuyu County[J].HEILONGJIANG AGRICULTURAL SCIENCES,2014,(01):150.
[6]郑 旭,赵秀梅,王连霞,等.25%阿维〖DK1〗·福〖DK1〗·莠锈悬浮种衣剂病害防效试验[J].黑龙江农业科学,2014,(05):54.
　ZHENG Xu,ZHAO Xiu-mei,WANG Lian-xia,et al.Field Efficacy Trial of 25% Abamectin·Thiram·Carboxin FSC Against Soybean Disease[J].HEILONGJIANG AGRICULTURAL SCIENCES,2014,(01):54.
[7]袁明. 齐齐哈尔大豆综合试验站现代农业产业示范基地建设成效的探讨[J].黑龙江农业科学,2013,(01):143.
　YUANMing.DiscussiononConstructionEffectofQiqiharSoybeanComprehensiveExperimentalStationofModernAgriculturalIndustrializationDemonstrationBase[J].HEILONGJIANG AGRICULTURAL SCIENCES,2013,(01):143.
[8]李亮,李泽宇,王丽娜,等.不同大豆品种耐盐性表现分析[J].黑龙江农业科学,2013,(02):17.
　LI Liang,LI Ze-yu,WANG Li-na,et al.Performance Analysis on Salt Tolerance for Different Soybean Varieties[J].HEILONGJIANG AGRICULTURAL SCIENCES,2013,(01):17.
[9]郑新利,张丙双,寇贺.东北地区大豆生产主要问题及发展对策[J].黑龙江农业科学,2012,(02):146.
　ZHENG Xin-li,ZHANG Bing-shuang,KOU He.Main Problems and Developing Countermeasuresof Soybean in Northeast China[J].HEILONGJIANG AGRICULTURAL SCIENCES,2012,(01):146.
[10]孙羽,冯延江,王麒,等.玉米茬原垄卡种大豆对其生长及产量的影响[J].黑龙江农业科学,2014,(03):26.
　SUN Yu,FENG Yan jiang,WANG Qi,et al.Effect of Original Ridge Tillage on the Growth and Yield of Soybean[J].HEILONGJIANG AGRICULTURAL SCIENCES,2014,(01):26.

备注/Memo

收稿日期：2021-09-22

第一作者:金勋(1962-),男，博士，高级农艺师，从事植物遗传育种研究。E-mail:jinxun6268@126.com。

更新日期/Last Update: 2022-04-01