JIANG Yao,CHEN Wenbo,CHEN Qibing.Cloning and Expression Characterization of Heat-resistant Gene [STBX]HSFA2[STBZ] in Wild Lily[J].Northern Horticulture,2022,(22):67-73.[doi:10.11937/bfyy.20220715]
野生湖北百合[STBX]HSFA2[STBZ]耐热基因的克隆及表达分析
- Title:
- Cloning and Expression Characterization of Heat-resistant Gene [STBX]HSFA2[STBZ] in Wild Lily
- 关键词:
- 野生百合; [STBX]HSFA2[STBZ]基因; 基因表达; 耐热性
- Keywords:
- wild lily; [STBX]HSFA2[STBZ] gene; genetic expression; heat-resistance
- 文献标志码:
- A
- 摘要:
- 以黔东南州野生百合叶片为试材，采用RACE及荧光定量PCR技术，研究了野生百合〖STBX〗HSFA2〖STBZ〗耐热基因的克隆及表达的影响，以期为黔东南州野生百合耐热能力提高及抗性改良提供参考依据。结果表明：获得1 435 bp 〖STBX〗HSFA2〖STBZ〗全长基因序列，该基因包含5′-UTR 77 bp、3′-UTR 155 bp和ORF 1 203 bp，可编码400个氨基酸，蛋白质分子量为45.3 kD。〖STBX〗LlHSFA2〖STBZ〗基因氨基酸序列与毛果杨、小叶杨同源性分别为87.56%和83.64%，表明其具有典型的热激转录因子结构域，DNA结合域在来源于不同物种中的HSF2A蛋白均比较保守。荧光定量结果表明，〖STBX〗LlHSFA2〖STBZ〗基因在根、鳞茎、叶组织里均有表达且存在差异，但在叶中的表达量明显高于根和鳞茎。经热激处理3～12 h，〖STBX〗HSFA2〖STBZ〗基因其表达量随着热激时间的增长呈上升趋势，但在24 h时其表达量下降但仍然高于对照。超量〖STBX〗HSFA2〖STBZ〗转化成功株系比野生型均有不同程度地超量表达，其中表达量最高的株系为OE-5和OE-7，对其进行热激处理后，其光化学淬灭系数、叶绿素含量均高于野生型对照，表现出更好的耐热性，这与模式植物拟南芥〖STBX〗HSFA2的结果类似，说明HSFA2〖STBZ〗在不同植株中均具有增强耐热性功能，是一个保守的植物适应高温的正调控因子。
- Abstract:
- Taking leaves of wild lily as test materials which from Southeast of Guizhou prefecture, the effects on cloning and expression characterization of heat-resistant gene 〖STBX〗HSFA2 〖STBZ〗in wild lily were studied by RACE and qRT-PCR method,in order to provide reference for improving the heat tolerance and resistance improvement of wild lily in Southeast of Guizhou.The results showed that the full length with 1 435 bp cDNA of 〖STBX〗HSFA2 〖STBZ〗gene was isolated 〖STBX〗HSFA2 〖STBZ〗gene,containing with 5′-untranslated region of 77 bp, 3′-untranslated region of 155 bp and open reading frame of 1 203 bp which encoding 400 amino acid with a molecular weight of 45.3 kD.This protein was highly homologous to know HSFA2,which sharing 87.56% and 83.64% identities with HSFA2 from Populus trichocarpa and Populus simonii respectively.The results showed that highly conserved DNA binding from the different species was in this protein.qRT-PCR analysis indicated that the expression leve l of HFA2 in root,bulbs and leaves was obviously different,but which in leaves of wild lily was significantly higher than that in root and bulbs.The relative expression level of 〖STBX〗HSFA2 〖STBZ〗gene was significantly elevated during 3—12 hours after heat treatment,but the expression level was down-regulated at 24 hours,which was higher than wild type.The relative expression levels of OE-5 and OE-7 transgenic lines which reaching the peak,were greater than that of non-transgenic plants.Photochemical quenching coefficient and chlorophyll content of transgenic plant were higher than that in non-transgenic plants after heat treatment.Furthermore,the results showed that the ability of transgene lines to heat stress was similar to that of Arabidopsis.This research obtained the different transgenic lines with higher resistance ability to heat stress,which was a highly conserved up-regulator to heat stress.
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备注/Memo
第一作者简介：蒋瑶（1982-），女，博士，教授，现主要从事植物生物技术等研究工作。E-mail:jiangyao0221@163.com.基金项目：贵州省科技厅自然科学基金资助项目（黔科合J字［2014］2153号）；贵州省科技厅自然科学基金重点资助项目（黔科合基础［2018］1413号）。收稿日期：2022-02-28