紅麻TIR1基因克隆及其表達(dá)載體構(gòu)建
發(fā)布時(shí)間：2018-08-17 20:11
【摘要】：【目的】分析紅麻運(yùn)輸抑制劑響應(yīng)蛋白1(TIR1)基因在不同組織和不同發(fā)育時(shí)期花藥中的表達(dá)情況,并構(gòu)建其超量表達(dá)載體和干擾載體,為研究該基因在雄蕊發(fā)育中的調(diào)控功能打下基礎(chǔ)�！痉椒ā扛鶕�(jù)紅麻花藥轉(zhuǎn)錄組數(shù)據(jù),利用生物信息學(xué)方法,克隆TIR1基因cDNA序列,實(shí)時(shí)熒光定量PCR(qPCR)分析TIR1基因在紅麻保持系722B和不育系722A根、莖、葉及不同發(fā)育時(shí)期花藥中的表達(dá)情況,并構(gòu)建超量表達(dá)載體和干擾載體�！窘Y(jié)果】TIR1基因的開放閱讀框(ORF)為1761 bp,編碼586個(gè)氨基酸(Gen Bank登錄號(hào)KY613992)。qPCR檢測(cè)結(jié)果顯示,與不育系722B相比,不育系722A TIR1基因在四分體期花藥中呈顯著下調(diào)表達(dá)(P0.05,下同),在莖和雙核期花藥中呈顯著上調(diào)表達(dá),在單核期花藥中極顯著上調(diào)表達(dá)(P0.01);而在根和葉中,保持系722B和不育系722A中TIR1基因表達(dá)水平差異均不顯著(P0.05)。超量表達(dá)載體p BI121-GFP-TIR1和干擾載體p ART27-p K-Tz-Tf構(gòu)建成功。【結(jié)論】紅麻TIR1基因編碼一個(gè)富含亮氨酸重復(fù)的F-box蛋白。紅麻不育系722A的單核期花藥TIR1基因表達(dá)較保持系722B極顯著上調(diào)表達(dá),可能與花藥敗育有關(guān)。構(gòu)建的超量表達(dá)載體和干擾載體,可用于紅麻TIR1基因功能及其與雄蕊發(fā)育的關(guān)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.
【作者單位】： 廣西大學(xué)農(nóng)學(xué)院;
【基金】：國(guó)家自然科學(xué)基金項(xiàng)目(31571719)
【分類號(hào)】：Q943.2;S563.5
本文編號(hào)：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.
【作者單位】： 廣西大學(xué)農(nóng)學(xué)院;
【基金】：國(guó)家自然科學(xué)基金項(xiàng)目(31571719)
【分類號(hào)】：Q943.2;S563.5
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