本文選題：銅綠假單胞菌 + 蛋白質(zhì)組學(xué)��； 參考：《廣西醫(yī)科大學(xué)》2007年碩士論文

【摘要】： 目的:找出銅綠假單胞菌野生型PAO1與黏液型PDO300菌株、PAO1lasI rhlI基因缺陷型菌株、PAO1 lasR rhlR基因缺陷型菌株之間的差異蛋白質(zhì),探討這些蛋白質(zhì)與銅綠假單胞菌生物膜形成的關(guān)系。 方法:(1)將同一份樣品配制成不同蛋白濃度的標本分別上樣檢測,對檢測所得蛋白質(zhì)圖譜進行分析。(2)根據(jù)最佳檢測蛋白濃度,將黏液型PDO300、野生型PAO1、PAO1 lasI rhlI基因缺陷型和PAO1 lasR rhlR基因缺陷型菌株的菌體蛋白分別采用CM10、IMAC3-Cu這兩種蛋白質(zhì)芯片,運用SELDI技術(shù)進行測定,篩選最佳蛋白質(zhì)芯片。(3)利用SELDI技術(shù)對黏液型PDO300、野生型PAO1、PAO1 lasI rhlI基因缺陷型和PAO1 lasR rhlR基因缺陷型菌株的菌體蛋白分別進行檢測,并采用Biomarker Wizard軟件進行分析。 結(jié)果:(1)檢測蛋白濃度為1.5μg/μl時,進行SELDI檢測可發(fā)現(xiàn)較多的蛋白峰。(2)同樣檢測蛋白濃度時,CM10芯片捕獲的蛋白峰數(shù)量比IMAC3-Cu芯片多。(3)銅綠假單胞菌野生型PAO1與黏液型PDO300菌株的菌體蛋白比較,有16個蛋白峰存在差異(P＜0.01)。與野生型PAO1菌株相比,PDO300菌株中有10個蛋白質(zhì)高表達,分子量分別為4,074Da、3,817Da、8,923Da、8,148Da、3,332Da、4,521Da、7,621Da、11,594Da、7,831Da和13,751Da;有6個蛋白質(zhì)低表達,分子量分別為6,199Da、6,899Da、12,379Da、6,115Da、6,763Da和8,576Da。(4)銅綠假單胞菌PAO1 lasI rhlI基因缺陷型與野生型PAO1菌株相比有1個蛋白質(zhì)低表達(P＜0.05),分子量為6,910Da。(5)銅綠假單胞菌PAO1 lasR rhlR基因缺陷型與野生型PAO1菌株之間有4個蛋白峰存在差異(P＜0.05)。在PAO1 lasR rhlR基因缺陷型菌株中有2個蛋白質(zhì)高表達,分子量分別為8,924Da和8,790Da:有2個蛋白質(zhì)低表達,分子量分別為6,735Da和6,986Da。 結(jié)論:(1)運用SELDI技術(shù)對細菌菌體蛋白進行分析的最佳檢測蛋白濃度為1.5μg/μl。 (2)與IMAC3-Cu芯片相比,CM10芯片更適于細菌菌體蛋白質(zhì)組學(xué)的研究。 (3)銅綠假單胞菌野生型PAO1與黏液型PDO300的菌體蛋白有16個蛋白質(zhì)表達明顯不同,這些差異蛋白可能與銅綠假單胞菌的表型和生物膜形成有關(guān)。 (4)銅綠假單胞菌野生型PAO1與PAO1 lasI rhlI基因缺陷型菌株之間有1個蛋白質(zhì)表達不同,野生型PAO1與PAO1 lasR rhlR基因缺陷型菌株之間有4個蛋白質(zhì)表達不同,這些蛋白質(zhì)可能與QS系統(tǒng)調(diào)控生物膜形成有關(guān)。 (5)用蛋白質(zhì)芯片和SELDI技術(shù)對銅綠假單胞菌進行蛋白質(zhì)組學(xué)研究,方法簡便、敏感性高,可發(fā)現(xiàn)一系列傳統(tǒng)研究手段難以測得的低分子量或低豐度的蛋白質(zhì)。
[Abstract]:Objective: to find out the differential proteins between wild-type PAO1 of Pseudomonas aeruginosa and PAO1lasI rhlI gene deficient strain of PAO1lasI rhlI gene, and to explore the relationship between these proteins and biofilm formation of Pseudomonas aeruginosa. Methods 1) the same sample was prepared into samples with different protein concentrations, and the protein profiles were analyzed. 2) according to the optimum protein concentration, The bacterial proteins of myxotypic PDO300, wild type PAO1, PAO1 lasI rhlI gene deficient type and PAO1 lasR rhlR gene deficient type strain were determined by SELDI technique by using CM10 / IMAC3-Cu protein chip, respectively. Screening the best protein microarray, using SELDI technique to detect myxotypic PDO300, wild type PAO1 PAO1 lasI rhlI gene deficient type and PAO1 lasR rhlR gene deficient type strain respectively, and to analyze them by Biomarker Wizard software. Results when the concentration of protein was 1.5 渭 g / 渭 l, The number of protein peaks captured by CM10 microarray was more than that of IMAC3-Cu chip by SELDI detection. There were 16 protein peaks of Pseudomonas aeruginosa wild-type PAO1 compared with myxic-type PDO300 strains (P < 0.01). Compared with wild-type PAO1 strains, 10 proteins were highly expressed, and their molecular weights were 4074 Da3817Dan8923Daan8148Da31332Dax4521Daf7621Da7594Da77831Da and 13751Da.There were 6 proteins that were low expressed. The molecular weight of the PAO1 lasI rhlI gene deficiency type of Pseudomonas aeruginosa was 6199 Daxin6899Daf12379Daji6115Da6763Da and 8576Da.Y4) there was a protein low expression between the PAO1 lasI rhlI gene deficient type of Pseudomonas aeruginosa and the wild type PAO1 strain (P < 0.05), and the molecular weight was 6910 Da. 5) between the PAO1 lasR rhlR gene deficiency type and the wild type PAO1 strain of Pseudomonas aeruginosa. There were four protein peaks (P < 0.05). Two proteins were highly expressed in PAO1 lasR rhlR gene deficient strains with molecular weight of 8924Da and 8790 Da, and two proteins were low expressed, with molecular weight of 6735Da and 6986 Da, respectively. ConclusionThe optimal concentration of SELDI for the analysis of bacterial protein is 1.5 渭 g / 渭 l. Compared with IMAC3-Cu chip, CM10 chip is more suitable for proteomics of bacteria. (3) the expression of 16 proteins in wild-type PAO1 and myxotypic PDO300 of Pseudomonas aeruginosa was significantly different, which might be related to the phenotype and biofilm formation of Pseudomonas aeruginosa. 1 protein expression was different between wild-type PAO1 and PAO1 lasI rhlI gene deficient strains of Pseudomonas aeruginosa, and 4 proteins were different between wild-type PAO1 and PAO1 lasR rhlR gene deficient strains. These proteins may be related to QS system regulating biofilm formation. The proteomics of Pseudomonas aeruginosa was studied by using protein chip and SELDI technique. The method was simple and sensitive. A series of low molecular weight or low abundance proteins could be found by traditional methods.
【學(xué)位授予單位】：廣西醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2007
【分類號】：R378

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