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厚藤脱水素基因IpDHN启动子IpDHN-Pro的克隆和调控转录活性分析
苏华香1,2, 郑洁旋1,2, 张会1,2, 何金实3, 简曙光1, 夏快飞1, 陈建通1, 张美1
1.中国科学院华南植物园, 广东省应用植物学重点实验室, 广州 510650;2.中国科学院大学, 北京 100049;3.海南大学热带农林学院, 海口 570228
摘要:  为了解厚藤(Ipomoea pes-caprae)脱水素基因IpDHN(GenBank登录号:KX426069)启动子的转录活性和对非生物胁迫和植物激素ABA的响应,通过染色体步移法克隆了IpDHN的上游启动子序列IpDHN-Pro,长度为974 bp。构建IpDHN-Pro调控下GUS转基因载体,转化拟南芥(Arabidopsis thaliana)植株获得IpDHN-Pro::GUS转基因植株并进行GUS染色,验证IpDHN-Pro启动转录活性以及在氯化钠、甘露醇、ABA处理后拟南芥GUS基因表达变化。结果表明,扩增获得的IpDHN-Pro序列包含多个顺式作用元件,包括1个ABRE、3个Myb转录因子结合位点、富含TC的重复序列以及Skn-1基序等。转基因拟南芥GUS染色及qRT-PCR表明该序列可驱动GUS基因在拟南芥稳定表达,且表达受高盐、渗透压及ABA的诱导。这表明IpDHN-Pro是一个盐旱、ABA诱导的启动子序列,可应用于相关的植物抗逆遗传工程研究。
关键词:  启动子  盐旱诱导  ABA诱导  脱水素  厚藤
DOI:10.11926/jtsb.4011
分类号:
基金项目:中国科学院A类战略性先导科技专项(XDA13020500);2018年中国科学院华南植物园大学生创新计划项目(67)资助
Cloning and Transcriptional Regulating Activities Analysis of A Dehydrin Promoter IpDHN-Pro from Ipomoea pes-caprae
SU Hua-xiang1,2, ZHENG Jie-xuan1,2, ZHANG Hui1,2, HE Jin-shi3, JIAN Shu-guang1, XIA Kuai-fei1, CHEN Jian-tong1, ZHANG Mei1
1.Provincial Key Laboratory of Applied Botany, South China Botanical Garden, Guangzhou 510650, China;2.University of Chinese Academy of Sciences, Beijing 100049, China;3.Institute of Tropical Agriculture and Forestry, Hainan University, Haikou 570228, China
Abstract:  Based on the previous research about Ipomoea pes-caprae cDNA library screening, a full length cDNA encoding dehydrin was characterized and named as IpDHN (GenBank accession:No. KX426069). In order to explore the transcriptional activity of the IpDHN promoter and the characteristics of this promoter region responding to abiotic stresses and ABA, the 5' upstream sequence of IpDHN (974 bp), named as IpDHN-Pro was cloned by genomic walking method, and then was subcloned into plant expression vector, in which GUS is under control of IpDHN-Pro. The transgenic Arabidopsis plants harboring IpDHN-Pro::GUS were checked by GUS staining assay, then the expression of GUS was further detected by qRT-PCR after the transgenic plants were challenged by NaCl, mannitol and ABA. The results showed that the IpDHN-Pro contained several cis-acting elements, including an ABRE, three Myb transcription factor binding sites, a TC-rich repeat, and a Skn-1 motif. The GUS staining and qRT-PCR showed that IpDHN-Pro could drive the stable expression of GUS gene in transgenic Arabidopsis, and this promoter was up-regulated by high salinity, osmotic stress and ABA treatment. Therefore, it was indicated that IpDHN-Pro was a salt/dehydration and ABA induced promoter sequence, and can be applied in plant genetic engineering research aiming at abiotic stress tolerance.
Key words:  Promoter  Salt/osmotic stress induced  ABA induced  Dehydrin  Ipomoea pes-caprae

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