【摘要】：前纖維蛋白(profilin,PRF)是一種植物中廣泛存在的微絲結合蛋白,對微絲動態(tài)重排進行雙向調控,在植物的各項生命活動中起非常重要的作用。為研究其在小麥(Triticum aestivum)中的功能,本研究以小麥光溫敏不育系BS366雄蕊的cDNA為模板,通過電子克隆和RT-PCR方法克隆獲得1個PRF類基因,并根據(jù)Gen Bank登錄順序命名為TaPRF7(Gen Bank No.KY940299)。利用生物信息學軟件對TaPRF7及其蛋白進行分析,結果表明,TaPRF7編碼區(qū)含有396個核苷酸,編碼131個氨基酸;其蛋白分子量約為14.2 KD,p I約為4.81,屬于穩(wěn)定蛋白;含有保守域PROF,有肌動蛋白(actin)、磷脂酰肌醇4,5-二磷酸(phosphatidyl inositol 4,5-diphosphate,PIP2)、多聚脯氨酸結合位點。TaPRF7基因與玉米(Zea mays)ZmPRF2、ZmPRF4、ZmPRF5,大麥(Hordeum vulgare)Hv PRF1,高粱(Sorghum bicolor)Sb PRF,水稻(Oryza sativa)Os PRF親緣關系較近,同源性也比較高。構建TaPRF7-16318h GFP融合蛋白表達載體,觀察其在擬南芥(Arabidopsis thaliana)原生質體亞細胞定位,顯示定位于細胞核和細胞質中;運用qRT-PCR分析小麥BS366不同組織及脫落酸(abscisic acid,ABA)、生長素(indoleaceticacid,IAA)、茉莉酸甲酯(methyl jasmonate,MeJA)、鹽(NaCl)、模擬干旱(PEG 6000)、水楊酸(salicylic acid,SA)、赤霉素(gibberellin,GA)、低溫(10℃)處理下TaPRF7表達特性。分析表明,TaPRF7在雄蕊中表達量最高,屬于生殖型表達。在ABA、PEG、GA、NaCl、IAA和低溫6種處理下,TaPRF7表達量都呈現(xiàn)出先升后降的趨勢,而在Me JA和SA處理下,TaPRF7的表達被顯著抑制。利用qRT-PCR分析該基因在不育系BS366和恢復系京411不育和可育環(huán)境下花藥發(fā)育3個時期的表達情況,結果表明,相比BS366,該基因在京411中3個時期表達量都很低,且在BS366不育環(huán)境下表達量明顯高于可育環(huán)境,隨著花藥發(fā)育時期的推進,在不育環(huán)境下該基因表達量呈上升趨勢。綜上結果推測TaPRF7可能參與了花藥開裂和低溫誘導不育等小麥分子信號轉導通路,為進一步研究TaPRF7基因在光溫敏小麥不育的分子機制提供了一定的理論依據(jù)。
[Abstract]:Prefibrin (profilin,PRF) is a widely existed microfilament binding protein in plants. It regulates the dynamic rearrangement of microfilaments in both directions and plays an important role in the life activities of plants. In order to study its function in wheat (Triticum aestivum), a PRF gene was obtained by electronic cloning and RT-PCR cloning using cDNA of wheat photo-thermosensitive sterile line BS366 stamen as template, and named TaPRF7 (Gen Bank No.KY940299 according to Gen Bank login sequence. TaPRF7 and its protein were analyzed by bioinformatics software. The results showed that the coding region of TaPRF7 contained 396 nucleotides encoding 131 amino acids, and its molecular weight was about 14.2 KD,p I about 4.81, which was a stable protein. The conserved domain PROF, contained actin (actin), phosphatidylinositol (actin), 5-diphosphatePIP2), and the polyproline binding site. The TaPRF7 gene was closely related to (Zea mays) ZmPRF2,ZmPRF4,ZmPRF5, barley (Hordeum vulgare) Hv PRF1, sorghum (Sorghum bicolor) Sb PRF, rice (Oryza sativa) Os PRF and had high homology. The expression vector of TaPRF7-16318h GFP fusion protein was constructed, and its localization in (Arabidopsis thaliana) protoplast subcellular of Arabidopsis thaliana was observed, which showed that it was located in the nucleus and cytoplasm of Arabidopsis thaliana. QRT-PCR was used to analyze the TaPRF7 expression characteristics of wheat BS366 treated with abscisic acid (abscisic acid,ABA), auxin (indoleaceticacid,IAA), methyl jasmonate (methyl jasmonate,MeJA), salt (NaCl), simulated drought (PEG 6000), salicylic acid (salicylic acid,SA), gibberellin (gibberellin,GA) and low temperature (10 鈩，

本文編號：2277700