本文選題：西瓜食酸菌 + RND家族外排轉(zhuǎn)運(yùn)體��； 參考：《新疆農(nóng)業(yè)大學(xué)》2015年碩士論文

【摘要】：西瓜食酸菌(Acidovorax citrulli)侵染葫蘆科植物會(huì)使其產(chǎn)生一種破壞力極大的瓜類細(xì)菌性果斑病(bacterial fruit blotch,BFB),該病害已在世界各國迅速傳播,對當(dāng)?shù)氐奈魈鸸袭a(chǎn)業(yè)造成嚴(yán)重的經(jīng)濟(jì)損失。目前銅素類殺菌劑廣泛應(yīng)用于防治瓜類細(xì)菌性果斑病,但過度使用,使得西瓜食酸菌抗銅能力逐漸增強(qiáng),因此,研究該病菌的抗銅機(jī)理顯得尤為突出。本研究首先對西瓜食酸菌RND(耐藥結(jié)節(jié)細(xì)胞分化)家族外排轉(zhuǎn)運(yùn)體的膜融合蛋白基因cusB進(jìn)行生物信息學(xué)分析,然后利用實(shí)驗(yàn)室保存的cusB基因缺失的突變體△cusB,通過比較野生型和突變體以及功能互補(bǔ)菌株在抗銅性、胞外分泌物檢測、致病性及過敏性壞死反應(yīng)等相關(guān)的生物學(xué)性狀之間的差異分析,研究cusB基因的功能,主要獲得以下結(jié)論:首先,cusB基因的生物信息學(xué)分析,cusB基因大小為1185bp,編碼具有394個(gè)氨基酸殘基的CusB蛋白亞基,主要作為一個(gè)轉(zhuǎn)運(yùn)體參與RND外排轉(zhuǎn)運(yùn)體的正常工作,西瓜食酸菌的CusB蛋白與大腸桿菌的CusB蛋白在理化性質(zhì)以及親緣關(guān)系上較近,預(yù)測該蛋白在1-30氨基酸殘基處疏水性較高,且存在一個(gè)跨膜結(jié)構(gòu)域,預(yù)測發(fā)現(xiàn)存在多個(gè)磷酸化位點(diǎn),分泌蛋白的亞細(xì)胞定位于周質(zhì)空間,其二級結(jié)構(gòu)存在α-螺旋、β-折疊、無規(guī)則卷曲。其次,cusB基因缺失突變體與野生型和功能互補(bǔ)菌株表型測定比較本研究利用雙親雜交的方法構(gòu)建功能互補(bǔ)菌株,與野生型相比,突變體在抗銅性,胞外多糖分泌,生物膜形成中顯著降低,而在功能互補(bǔ)菌株中這些差異明顯減小,表明西瓜食酸菌的cusB基因與這3種表型聯(lián)系緊密,但cusB基因的缺失沒有影響致病性及過敏性壞死反應(yīng)、胞外纖維素酶的分泌、胞外蛋白酶的分泌,表明該基因與這3種表型關(guān)系不密切。
[Abstract]:The infection of Acidovorax citrulliae with Cucurbitaceae will produce a kind of bacterial fruit blotchchus bFGF, which has been spreading rapidly in many countries all over the world, and has caused serious economic losses to the local muskmelon industry. At present, copper fungicides are widely used to control bacterial fruit spot disease of melon, but the copper resistance ability of watermelon acid-eating bacteria is gradually enhanced by excessive use. Therefore, the study of copper resistance mechanism of this pathogen is particularly prominent. In this study, the membrane fusion protein gene cusB of the efflux transporter of RND (resistant nodular cell differentiation) family in watermelon was analyzed by bioinformatics. Then we compared the copper resistance and extracellular secretion of wild-type mutants and functional complementary strains by using the mutant cusB gene deleted in laboratory. To study the function of cusB gene by analyzing the differences of biological characters such as pathogenicity and anaphylactic necrosis reaction. The main conclusions are as follows: firstly, the bioinformatics analysis of CusB gene is 1185 BP, encoding 394 amino acid residues of CusB protein subunit, mainly as a transporter to participate in the normal work of RND efflux transporter. The relationship between CusB protein and Escherichia coli CusB protein was close in physicochemical properties and phylogenetic relationship. It was predicted that the protein was hydrophobic at 1-30 amino acid residues and had a transmembrane domain. It was predicted that there were many phosphorylation sites. The subcell of secretory protein is located in the periplasmic space, and its secondary structure is 偽 -helix, 尾 -fold and irregular curl. Secondly, the phenotypic determination of CusB gene deletion mutants was compared with wild type and functional complementary strains. In this study, we used the method of parental hybridization to construct functional complementary strains. Compared with wild type, the mutants were copper resistant and exopolysaccharide secreted. These differences were significantly decreased in biofilm formation and decreased in functional complementary strains, indicating that the cusB gene was closely related to the three phenotypes, but the absence of cusB gene had no effect on pathogenicity and anaphylactic necrosis. The secretion of extracellular cellulase and extracellular protease showed that the gene was not closely related to the three phenotypes.
【學(xué)位授予單位】：新疆農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：S432.4

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本文編號：2020504