本文選題：聚球藻7002　切入點(diǎn)：LamA基因　出處：《中國(guó)地質(zhì)大學(xué)(北京)》2017年博士論文　論文類型：學(xué)位論文

【摘要】：礦石燃料燃燒排放到大氣中的CO_2對(duì)全球氣候的變化起了決定性作用。由于藍(lán)藻具有生長(zhǎng)速度快、CO_2固定速率高、其生物質(zhì)和細(xì)胞產(chǎn)物具有較高的附加值等特點(diǎn),利用藍(lán)藻生物固碳具有突出的應(yīng)用價(jià)值。本研究以藍(lán)藻為受體,將海棲熱胞菌Msb8中合成海帶多糖酶的LamA基因轉(zhuǎn)入聚球藻7002中。實(shí)現(xiàn)了利用密閉型光生物反應(yīng)器培養(yǎng)藍(lán)藻固碳和生產(chǎn)高附加值重組蛋白的雙重目的。本文在成功構(gòu)建了轉(zhuǎn)LamA基因的重組聚球藻的基礎(chǔ)上,對(duì)重組聚球藻生產(chǎn)的LamA編碼的海帶多糖酶的酶活性和理化特性進(jìn)行了研究。結(jié)果表明,重組海帶多糖酶的最大相對(duì)酶活性為4.325 U?mg-1 dry mass。其最適酶反應(yīng)條件為pH=8.0,70℃。重組海帶多糖酶在pH值為6-8的范圍內(nèi)具有較高的活性,同時(shí)表現(xiàn)出很強(qiáng)的耐熱性。在80℃和90℃其酶活性的半衰期分別為40 min和20 min。根據(jù)密閉型光生物反應(yīng)器的優(yōu)缺點(diǎn)設(shè)計(jì)了改良型攪拌罐式光生物反應(yīng)器。通過(guò)空載試驗(yàn)和完全析因設(shè)計(jì),對(duì)反應(yīng)器的動(dòng)力學(xué)特性和重組聚球藻7002的生長(zhǎng)特性進(jìn)行分析,確定了對(duì)重組聚球藻Xmax的主要影響因素依次為:光照強(qiáng)度CO_2濃度溫度*溫度溫度CO_2濃度*光照強(qiáng)度;對(duì)μmax的主要影響因素依次為:CO_2濃度光照強(qiáng)度溫度。反應(yīng)器的最佳運(yùn)行條件為進(jìn)氣量為0.5 L·min-1,進(jìn)氣CO_2濃度為5%,光照強(qiáng)度為300μmol·m-2·s-1,溫度為30℃。Xmax和μmax分別達(dá)到3.929 g·L-1和0.015 h-1。此外,當(dāng)光照周期由連續(xù)光照變?yōu)殚g隙光照,重組聚球藻的比生長(zhǎng)速率、產(chǎn)率、CO_2固定速率和最大生物質(zhì)濃度均下降。高濃度CO_2(5%)對(duì)重組聚球藻7002的生長(zhǎng)和海帶多糖酶的相對(duì)活性均具抑制作用。雖然重組聚球藻在進(jìn)氣CO_2濃度為15%時(shí)仍可生長(zhǎng),但其Xmax、μmax和海帶多糖酶相對(duì)酶活性分別為最適條件下的36.9%、40.7%和75.1%。通過(guò)pH控制和高濃度CO_2馴化培養(yǎng)的方法均可提高重組聚球藻7002的Xmax和μmax。結(jié)果顯示,加入碳酸-碳酸氫鈉緩沖液效果最佳,比未加入緩沖液的情況下(2.507 g·L-1)Xmax和μmax分別提高了33.3%和15.8%。但由于pH值的升高,進(jìn)氣CO_2的固定效率降低,高濃度CO_2馴化的重組聚球藻7002初代Xmax和μmax比控制組提高了24.4%和11.2%。并且數(shù)代培養(yǎng)后,其適應(yīng)性穩(wěn)中有升。
[Abstract]:The CO_2 released from the combustion of mineral fuels into the atmosphere plays a decisive role in global climate change. Because cyanobacteria have the characteristics of fast growth rate and high fixation rate of COSP, their biomass and cell products have higher added value, etc. Biological carbon sequestration by cyanobacteria is of great application value. In this study, cyanobacteria were used as receptors. The LamA gene of synthesizing kelp polysaccharide enzyme from Msb8 of Pyrocystis martensii was transferred into Chlorella 7002. The purpose of carbon sequestration of cyanobacteria and production of high value-added recombinant protein were achieved by using closed photobioreactor. On the basis of the recombinant Chlorella sp. LamA gene, The enzyme activity and physical and chemical properties of the LamA encoded kelp polysaccharide enzyme produced by recombinant Chlorella spp were studied. The results showed that the maximum relative enzyme activity of the recombinant kelp polysaccharide enzyme was 4.325 U? The optimum reaction conditions of mg-1 dry were as follows: pH = 8.0 ~ 70 鈩，

本文編號(hào)：1575560