本文選題：hrp基因 + HrpA蛋白�。� 參考：《福建農(nóng)林大學(xué)》2008年碩士論文

【摘要】： Hrp(hypersensitive response and pathogenicity,Hrp)基因廣泛存在于植物病原細(xì)菌中,參與Hrp極毛系統(tǒng)的組裝,不僅可運(yùn)輸效應(yīng)蛋白至植物細(xì)胞,導(dǎo)致HR(過敏性反應(yīng))或抗病性,還可運(yùn)輸致病性和毒性蛋白或抑制子活性物質(zhì),引起植物病害。hrp基因多串聯(lián)成簇且相當(dāng)保守,在誘導(dǎo)培養(yǎng)基或植物體內(nèi),基因簇中的hrpA基因編碼細(xì)菌表面附屬物-Hrp極毛的結(jié)構(gòu)蛋白HrpA。Hrp極毛在植物-病原細(xì)菌互作中扮演十分重要的角色。病原細(xì)菌能否使親和性植物發(fā)病與其是否有Hrp極毛有直接的關(guān)系。 在對hrp基因進(jìn)行深入研究的基礎(chǔ)上,我們嘗試設(shè)計(jì)新的化學(xué)藥劑和抗體阻止HrpⅢ型分泌系統(tǒng)的組裝或致病毒力蛋白因子的分泌運(yùn)輸,從而有效限制植物細(xì)菌病害的發(fā)展。本研究以丁香假單胞菌番茄變種DC3000(Pseudomonassyringae pv.tomato DC3000)hrp基因表達(dá)的工程蛋白HrpA/Ps作為抗原,免疫BALB/c小鼠,制備單克隆雜交瘤細(xì)胞抗體,然后從雜交瘤細(xì)胞系中提取RNA,反轉(zhuǎn)錄合成cDNA,并以此為模板,PCR擴(kuò)增獲得全套抗體的可變區(qū)基因,利用從PQY1.5質(zhì)粒擴(kuò)增出一柔性肽段(Linker)連接成基因工程單鏈抗體(single chainvariable fragment,scFv),再通過基因工程方法,將scFv與噬菌粒pCANTAB-5E連接,轉(zhuǎn)化大腸桿菌TG1菌株,獲得單鏈抗體噬菌體文庫,以便利用噬菌體展示技術(shù)對抗體庫進(jìn)行淘選,獲得了HrpA特異性的單鏈抗體。 以單克隆抗體為來源的單鏈抗體,親和力和特異性都較好,可以為進(jìn)一步進(jìn)行植物抗病基因工程和植物抗體研究提供基因來源。
[Abstract]:The Hrp(hypersensitive response and pathogenicity HRP gene is widely present in plant pathogenic bacteria and participates in the assembly of Hrp polar hair system, which can not only transport effector protein to plant cells, but also lead to HR-allergic reaction or disease resistance. It can also transport pathogenicity and toxic protein or suppressor active substances, causing plant disease hrp genes to cluster and quite conserved, in induction medium or plant body, The hrpA gene in the gene cluster encodes the structural protein HrpA.Hrp polar hair, a bacterial surface appendage, which plays a very important role in the plant-pathogenic bacteria interaction. Whether pathogenic bacteria can cause affinity plant disease is directly related to the presence of Hrp hair. Based on the in-depth study of hrp gene, we try to design new chemical agents and antibodies to prevent the assembly of Hrp 鈪，

本文編號：1870248