本文選題：芽孢桿菌 + 炭疽　； 參考：《軍事醫(yī)學(xué)》2017年03期

【摘要】：目的構(gòu)建炭疽芽孢桿菌sigF基因缺失株,以及缺失株的sigF回復(fù)株,分析缺失株、回復(fù)株表型特點(diǎn)并明確sigF基因?qū)ρ挎咝纬傻挠绊?為后續(xù)研究提供基礎(chǔ)。方法采用同源重組技術(shù)在炭疽菌A16D2(pXO1+pXO2~-)菌株sigF基因處插入大觀(guān)霉素抗性基因,取代sigF基因,構(gòu)建了sigF基因缺失突變株。通過(guò)構(gòu)建回復(fù)株質(zhì)粒并電轉(zhuǎn)到sigF基因缺失株中,構(gòu)建了sigF基因的回復(fù)株。通過(guò)生長(zhǎng)曲線(xiàn)測(cè)定、糖代謝能力比較、顯微鏡觀(guān)察芽孢形成及微流控實(shí)驗(yàn)實(shí)時(shí)觀(guān)察細(xì)胞生長(zhǎng)等方法分析突變株和回復(fù)株的特點(diǎn)。結(jié)果獲得炭疽菌A16D2菌株sigF基因缺失突變株和相應(yīng)的回復(fù)株。生長(zhǎng)曲線(xiàn)測(cè)定表明,突變株較出發(fā)菌株在繁殖體階段生長(zhǎng)狀態(tài)無(wú)明顯差異;糖代謝實(shí)驗(yàn)表明,突變株與出發(fā)菌株在對(duì)碳水化合物的利用上差異不顯著;顯微鏡觀(guān)察顯示,突變株喪失了形成芽孢的能力、回復(fù)株恢復(fù)了部分形成芽孢的能力;微流控實(shí)驗(yàn)觀(guān)察顯示,缺失株細(xì)胞保持在不對(duì)稱(chēng)分裂的狀態(tài),回復(fù)株能形成芽孢。結(jié)論成功獲得了炭疽菌A16D2sigF基因缺失突變株。該研究證明sigF基因?yàn)樘烤揖纬裳挎咚匦?但不是細(xì)胞生長(zhǎng)所必需的因子。
[Abstract]:Objective to construct the sigF gene deletion strain of Bacillus anthracis and the sigF recovery strain of the deletion strain, to analyze the phenotypic characteristics of the deleted strain and to clarify the effect of sigF gene on the spores formation, and to provide a basis for further study. Methods the gene of spectinomycin resistance was inserted into the sigF gene of anthrax strain A16D2(pXO1 pXO _ 2 by homologous recombination technique, replacing the sigF gene, and the mutant of sigF gene deletion was constructed. By constructing the plasmid of the sigF gene and transferring it to the sigF gene deletion strain, the sigF gene recovery strain was constructed. The characteristics of mutants and recovers were analyzed by means of growth curve determination, comparison of glucose metabolism ability, observation of spores formation by microscope and real-time observation of cell growth by microfluidic experiments. Results the sigF gene deletion mutants and corresponding recovers of anthrax A16D2 strain were obtained. The growth curve test showed that there was no significant difference in the growth status of the mutant strain compared with the original strain in the reproductive phase, the glucose metabolism test showed that there was no significant difference in the utilization of carbohydrate between the mutant strain and the original strain, and the microscopic observation showed that there was no significant difference between the two strains in the utilization of carbohydrates. The mutant lost the ability to form spores, and the recovery strain recovered part of the ability to form spores, and the microfluidic experiments showed that the cells of the missing strain remained in the state of asymmetric division, and the recovered strain could form spores. Conclusion the mutant of A16D2sigF gene deletion of anthrax was successfully obtained. This study demonstrated that the sigF group is necessary for the formation of spores by anthrax, but is not an essential factor for cell growth.
【作者單位】： 上海海洋大學(xué)食品學(xué)院;軍事醫(yī)學(xué)科學(xué)院生物工程研究所病原微生物生物安全國(guó)家重點(diǎn)實(shí)驗(yàn)室;
【基金】：國(guó)家自然科學(xué)基金資助項(xiàng)目(81271785)
【分類(lèi)號(hào)】：R378.72

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