红麻TIR1基因克隆及其表达载体构建
发布时间：2018-08-17 20:11
【摘要】：【目的】分析红麻运输抑制剂响应蛋白1(TIR1)基因在不同组织和不同发育时期花药中的表达情况,并构建其超量表达载体和干扰载体,为研究该基因在雄蕊发育中的调控功能打下基础。【方法】根据红麻花药转录组数据,利用生物信息学方法,克隆TIR1基因cDNA序列,实时荧光定量PCR(qPCR)分析TIR1基因在红麻保持系722B和不育系722A根、茎、叶及不同发育时期花药中的表达情况,并构建超量表达载体和干扰载体。【结果】TIR1基因的开放阅读框(ORF)为1761 bp,编码586个氨基酸(Gen Bank登录号KY613992)。qPCR检测结果显示,与不育系722B相比,不育系722A TIR1基因在四分体期花药中呈显著下调表达(P0.05,下同),在茎和双核期花药中呈显著上调表达,在单核期花药中极显著上调表达(P0.01);而在根和叶中,保持系722B和不育系722A中TIR1基因表达水平差异均不显著(P0.05)。超量表达载体p BI121-GFP-TIR1和干扰载体p ART27-p K-Tz-Tf构建成功。【结论】红麻TIR1基因编码一个富含亮氨酸重复的F-box蛋白。红麻不育系722A的单核期花药TIR1基因表达较保持系722B极显著上调表达,可能与花药败育有关。构建的超量表达载体和干扰载体,可用于红麻TIR1基因功能及其与雄蕊发育的关系研究。
[Abstract]:[objective] to analyze the expression of kenaf transport inhibitor response protein 1 (TIR1) gene in anthers of different tissues and different developmental stages, and to construct its overexpression vector and interference vector. [methods] based on the transcriptional data of kenaf anther, the cDNA sequence of TIR1 gene was cloned by bioinformatics. The expression of TIR1 gene in root, stem, leaf and anther of kenaf maintainer line 722B and sterile line 722A was analyzed by real-time fluorescence quantitative PCR (qPCR). [results] the open reading frame (ORF) of TIR1 gene was 1761 BP, encoding 586 amino acid (Gen Bank accession number KY613992) .qPCR analysis showed that compared with sterile line 722B, the open reading frame (ORF) of TIR1 gene was 1761 BP, encoding 586 amino acid (Gen Bank login number KY613992). The expression of 722A TIR1 gene was significantly down-regulated in tetrad anthers (P0.05, the same below), significantly up-regulated in stem and binuclear anthers, and very significantly up-regulated in mononuclear anthers (P0.01), but in roots and leaves. There was no significant difference in TIR1 gene expression between maintainer line 722B and sterile line 722A (P0.05). The overexpression vector p BI121-GFP-TIR1 and interference vector p ART27-p K-Tz-Tf were successfully constructed. [conclusion] kenaf TIR1 gene encodes a Leucine-rich repeat F-box protein. The expression of TIR1 gene in anther of kenaf male sterile line 722A was significantly higher than that of maintainer line 722B, which might be related to anther abortion. The constructed overexpression vector and interference vector can be used to study the function of kenaf TIR1 gene and its relationship with stamen development.
【作者单位】： 广西大学农学院;
【基金】：国家自然科学基金项目(31571719)
【分类号】：Q943.2;S563.5
本文编号：2188768
[Abstract]:[objective] to analyze the expression of kenaf transport inhibitor response protein 1 (TIR1) gene in anthers of different tissues and different developmental stages, and to construct its overexpression vector and interference vector. [methods] based on the transcriptional data of kenaf anther, the cDNA sequence of TIR1 gene was cloned by bioinformatics. The expression of TIR1 gene in root, stem, leaf and anther of kenaf maintainer line 722B and sterile line 722A was analyzed by real-time fluorescence quantitative PCR (qPCR). [results] the open reading frame (ORF) of TIR1 gene was 1761 BP, encoding 586 amino acid (Gen Bank accession number KY613992) .qPCR analysis showed that compared with sterile line 722B, the open reading frame (ORF) of TIR1 gene was 1761 BP, encoding 586 amino acid (Gen Bank login number KY613992). The expression of 722A TIR1 gene was significantly down-regulated in tetrad anthers (P0.05, the same below), significantly up-regulated in stem and binuclear anthers, and very significantly up-regulated in mononuclear anthers (P0.01), but in roots and leaves. There was no significant difference in TIR1 gene expression between maintainer line 722B and sterile line 722A (P0.05). The overexpression vector p BI121-GFP-TIR1 and interference vector p ART27-p K-Tz-Tf were successfully constructed. [conclusion] kenaf TIR1 gene encodes a Leucine-rich repeat F-box protein. The expression of TIR1 gene in anther of kenaf male sterile line 722A was significantly higher than that of maintainer line 722B, which might be related to anther abortion. The constructed overexpression vector and interference vector can be used to study the function of kenaf TIR1 gene and its relationship with stamen development.
【作者单位】： 广西大学农学院;
【基金】：国家自然科学基金项目(31571719)
【分类号】：Q943.2;S563.5
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