本文選題：木葡糖酸醋桿菌 + 細菌纖維素��； 參考：《天津科技大學》2017年碩士論文

【摘要】：近年來,由于細菌纖維素所具有的高純度,高親水性、高生物相容性以及良好的機械性能被廣泛應用于生物醫(yī)藥行業(yè),造紙行業(yè)以及食品等行業(yè)。木葡糖酸醋桿菌Gluconacetobacter xyinus作為細菌纖維素的模式生產(chǎn)菌株,其生產(chǎn)細菌纖維素的確切機制尚不明確。本實驗室之前已證實在G.xylinus CGMCC 2955中存在類似費氏弧菌的LuxI/LuxR型群體感應系統(tǒng)。大量文獻表明群體感應與生物膜的合成有密切的關系,此本文針對群體感應與細菌纖維素合成之間的相關性進行了探究。首先利用分子生物學手段構建群體感應關鍵基因luxR過表達載體,將其電轉(zhuǎn)化到G.xylinus,通過靜置培養(yǎng)和振蕩培養(yǎng)兩種培養(yǎng)方式來比較原始菌株與過表達菌株發(fā)酵性能的差異,并比較細菌纖維素的結構性能差異。結果發(fā)現(xiàn),在振蕩培養(yǎng)條件下,與G.x相比,G.x-pMV24-luxR的菌體生長速度明顯減緩,到發(fā)酵結束,菌體量降低15.51%,葡萄糖酸的產(chǎn)量提高了 12.11%,對葡萄糖酸的利用率提高了 20.18%,但二者總酸產(chǎn)量無明顯差異。在靜置條件下,與G.x相比,G.x-pMV24-luxR的纖維素產(chǎn)量提高了 36.45%,且在16d葡萄糖酸含量降低了23.68%。對比各菌株所產(chǎn)的細菌纖維素的性能,傅里葉紅外光譜(Fourier Transform infrared spectroscopy, FT-IR)分析由于差異均存在于非結晶區(qū),表明細菌纖維素內(nèi)官能團結構沒有明顯差異;X射線衍射(X-ray diffraction.,XRD)表明G.x-pMV24-luxR產(chǎn)的纖維素結晶度明顯降低;TG/DTG表明G.x-pMV24-luxR產(chǎn)纖維素的熱穩(wěn)定性降低。利用代謝組學方法,對G.x以及G.x-pMV24-luxR胞內(nèi)小分子代謝物的的差異進行分析,探究群體感應對胞內(nèi)代謝的影響。結果表明luxR過表達對中心代謝、氨基酸代謝、脂代謝均有影響。G.x-pMV24-luxR大量合成葡萄糖酸,并且對嘌呤嘧啶的從頭合成也具有促進作用,但是卻抑制了氨基酸代謝和脂代謝,海藻糖作為一種細胞保護性物質(zhì),在葡萄糖酸大量合成的同時其含量也明顯增加。
[Abstract]:In recent years, bacterial cellulose has been widely used in biomedical industry, paper industry and food industry because of its high purity, high hydrophilicity, high biocompatibility and good mechanical properties. As a model strain of bacterial cellulose production, the exact mechanism of the production of bacterial cellulose by Gluconacetobacter xyinus is unclear. It has been previously confirmed in our laboratory that there is a LuxI/LuxR type population sensing system in G.xylinus CGMCC 2955 similar to Vibrio flexneri. A large number of literatures have shown that there is a close relationship between population induction and biosynthesis of biofilm. The correlation between group induction and bacterial cellulose synthesis is explored in this paper. Firstly, the overexpression vector of luxR gene was constructed by molecular biology, and transformed into G. xylinus. The fermentation performance of the original strain was compared with that of the overexpression strain by static culture and oscillatory culture. The structure and properties of bacterial cellulose were compared. The results showed that the growth rate of G. x-pMV24-luxR was significantly slower than that of G.x under oscillatory culture conditions, and the growth rate of G. x-pMV24-luxR was significantly slower than that of G.x, until the end of fermentation. The yield of gluconic acid and the utilization rate of gluconic acid were increased by 15.51 and 12.11 respectively, but there was no significant difference between them. Under static conditions, the cellulose yield of G.x-pMV24-luxR increased by 36.45, and the gluconic acid content decreased by 23.68% at 16 days compared with G. x. Comparing the properties of bacterial cellulose produced by each strain, Fourier Transform infrared spectroscopy (FT-IRI) analysis showed that the differences existed in the amorphous region. The results showed that there was no obvious difference in the structure of functional groups in bacterial cellulose. X-ray diffraction (XRD) showed that the crystallinity of cellulose produced by G.x-pMV24-luxR was obviously decreased. The results showed that the thermal stability of cellulose produced by G.x-pMV24-luxR was lower than that of TG / DTG. In order to explore the effect of population induction on intracellular metabolism, the differences between G. x and G.x-pMV24-luxR small molecule metabolites were analyzed by means of metabonomics. The results showed that the overexpression of luxR had an effect on the metabolism of central, amino acid and lipid. G.x-pMV24-luxR also promoted the ab initio synthesis of purine pyrimidine, but inhibited the metabolism of amino acid and lipid. Trehalose, as a cytoprotective substance, increases the content of gluconic acid in a large amount at the same time.
【學位授予單位】：天津科技大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：Q78;O636.11

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