青稞HVA1和blt4.9基因对模拟水分胁迫的响应差异及其在抗旱育种中的应用  

Difference of HVA1 and blt4.9 Gene Expression Patterns under Simulated Drought Stress and the Potention Application in Drought Tolerance Breeding in Hulless Barley

作　　者：王越 姚晓华[1,2] 吴昆仑 白羿雄[1,2] 魏晓星 WANG Yue;YAO Xiaohua;WU Kunlun;BAI Yixiong;WEI Xiaoxing(Qinghai University,Xining,Qinghai 810016,China;Qinghai Key Laboratory of Hulless Barley Geneticsand Breeding/Qinghai Subcenter of National Hulless Barley Improvement,Xining,Qinghai 810016,China;College of Eco-Environmental Engineering,Qinghai University,Xining,Qinghai 810016,China)

机构地区：[1]青海大学,青海西宁810016 [2]青海省青稞遗传育种重点实验室/国家麦类改良中心青海青稞分中心,青海西宁810016 [3]青海大学生态环境工程学院,青海西宁810016

出　　处：《麦类作物学报》2019年第6期666-674,共9页Journal of Triticeae Crops

基　　金：国家自然科学基金项目(31660388;31160284);现代农业产业技术体系建设专项(CARS-05);青海省科技厅项目(2019-ZJ-7075;2017-NK-A7)

摘　　要：为了解青稞HVA1和blt4.9基因在抗旱方面的作用及其差异,以青稞品种昆仑12号为材料,利用RT-PCR从8个候选内参基因(DHN1、GAPDH、Actin-1、Actin-2、18SrRNA-1、18SrRNA-2、TC139057和PKABA)中筛选出在15%PEG模拟水分胁迫下表达稳定的基因,同时以筛选到的稳定表达的基因为参照,采用qRT-PCR技术研究青稞HVA1基因和blt4.9基因在模拟水分胁迫条件下的表达差异及在不同抗旱性青稞品种鉴定中的应用。结果表明:(1)筛选到1个在干旱胁迫下青稞叶片中能稳定表达的内参基因TC139057;(2)在1%~30%PEG模拟干旱胁迫下,随着胁迫时间的延长,青稞HVA1和blt4.9基因表达量呈先增后降趋势,25%PEG处理下表达量最高,且在较低浓度PEG处理下基因表达量为HVA1>blt4.9,而在较高浓度PEG处理下反之;(3)15%PEG处理1~144h,两基因表达量也呈先增后降趋势,胁迫48h后HVA1基因表达量最高,胁迫96h后blt4.9基因表达量最高,且处理前期基因表达量为HVA1>blt4.9,后期反之;(4)1~500μmol·L^-1 ABA处理下,两基因表达量仍呈先增后降趋势,HVA1基因比blt4.9基因敏感,且在较低ABA浓度下基因表达量为HVA1>blt4.9,在较高浓度下反之;(5)获得转青稞HVA1和blt4.9基因拟南芥植株,其主要抗旱性生理指标优于野生型;且在较轻胁迫下,转HVA1植株的生理指标优于转blt4.9植株,而在较重胁迫处理下情况相反;(6)在模拟水分胁迫下,青稞叶片中两基因的表达量,抗旱性强的显著高于抗旱性弱的材料(P<0.01),且较轻胁迫下HVA1基因较敏感,反之blt4.9基因敏感。本研究结果为HVA1和blt4.9基因在青稞抗旱育种中的应用奠定了基础。In order to study the difference roles of drought resistance genes HVA1and blt4.9in hulless barley under drought,eight candidate internal reference genes (DHN1,GAPDH,actin-1,actin-2, 18SrRNA-1,18SrRNA-2,TC139057and PKABA)were screened in Kunlun 12under 15% PEG.The stable reference gene was used to study the differences of HVA1and blt4.9 gene expression in hulless barley under simulated drought stress and their application in the identification of different drought-resistance cultivars.The results showed that:a stable reference gene TC139057 was selected undersimulated drought stress in hulless barley;using TC139057as the reference gene,the expression levels of HVA1and blt4.9 genes in hulless barley were higher than those of the control(P<0.01)under the simulated drought stress,which showed a change trend of first increasing and then decreasing.The highest expression levels of these two genes appeared under 25% PEG treatment.Under lower concentration of PEG treatment(1%-20%),the expression level of HVA1 was higher than that of blt4.9,but under the higher concentration of PEG (25%-30%),the expression level of blt4.9 was higher.The two genes stably expressed from 1to 144hunder 15% PEG treatment,and the expression level increased first and then decreased with the prolonging of stress time.The peak of expression level for HVA1 and blt4.9 appeared at 48hand 96h,respectively.The gene expression level of HVA1 was higher than that of blt4.9from 1hto 48h,which is in contrast to that from 96hto 144h. Under the ABA treatment(1-500μmol·L^-1),the expression levels of the two genes also showed a change trend of first increasing and then decreasing.The highest expression level of HVA1 gene appeared under 10μmol·L^-1 ABA,but that of blt4.9 gene appeared under 50μmol·L^-1 ABA,which was significantly different from the control(P<0.01).The expression level of HVA1 was higher than that of blt4.9 under 1-10mmol·L^-1 ABA,in contrast to that under 50-500mmol·L^-1 ABA.The transgenic HVA1and blt4.9 Arabidopsis plants were achieved

关 键 词：青稞 HVA1基因 blt4.9基因 内参基因 实时荧光定量PCR 

分 类 号：S512.3[农业科学—作物学] S330