本文選題：綠色木霉 + 液態(tài)發(fā)酵��； 參考：《甘肅農(nóng)業(yè)大學(xué)》2015年碩士論文

【摘要】：本論文以實驗室保存的產(chǎn)纖維素酶菌株綠色木霉(Trichoderma viride)L4C為試驗材料,采用單因素試驗和正交試驗方法優(yōu)化該菌株液體發(fā)酵產(chǎn)酶條件;采用紫外誘變、亞硝基胍誘變及復(fù)合誘變方法,經(jīng)過纖維素水解圈的初篩及DNS法測酶活選育出產(chǎn)酶活力較高且遺傳性穩(wěn)定的菌株,結(jié)果如下:1、通過對菌株綠色木霉L4C液體產(chǎn)酶發(fā)酵條件進行單因素的優(yōu)化試驗,得到產(chǎn)酶培養(yǎng)的優(yōu)化條件:稻草和纖維素粉復(fù)合物為最佳碳源,硫酸銨為最適氮源,接種量為10%,培養(yǎng)時間為4 d,培養(yǎng)溫度為28℃,培養(yǎng)基初始pH為4.5。在單因素的基礎(chǔ)上,通過多因素正交試驗得到最佳產(chǎn)酶條件:接種量為15%,培養(yǎng)時間為5 d,培養(yǎng)溫度為28℃,初始pH為5.0。在最佳產(chǎn)酶條件下,CMCase為921.56U/mL,FPA為745.02 U/mL,半纖維素酶活為556.71 U/mL,分別比優(yōu)化前基礎(chǔ)培養(yǎng)基產(chǎn)酶提高了56.7%,65.0%和66.8%。2、通過紫外線(UV)誘變試驗,明確了紫外誘變的適宜條件:15 w紫外燈,距離30 cm,照射處理時間300 s,孢子懸浮液稀釋度為10-6。此條件下的誘變效應(yīng)為:致死率為85.0%。誘變后,獲得一株高產(chǎn)纖維素酶菌株-綠色木霉Z15(Trichoderma viride.Z15)。搖瓶發(fā)酵后,CMCase為1196.21 U/mL,FPA為863.56U/mL,其產(chǎn)酶能力分別較出發(fā)菌株提高了1.30倍和1.16倍。傳代試驗證明,該菌株具有較好的遺傳穩(wěn)定性。3、通過亞硝基胍(NTG)誘變試驗,明確了NTG誘變的適宜條件:NTG 0.6mg/mL,處理時間為50 min。該條件下的誘變效應(yīng)為:孢子致死率為80.4%,正突變率8.01%。誘變后,獲得一株高產(chǎn)纖維素酶菌株-綠色木霉N12(Trichoderma viride.N12)。搖瓶發(fā)酵后,CMCase為1410.02 U/mL,FPA為992.67U/mL,其產(chǎn)酶能力較出發(fā)菌株提高了18.5%和9.7%。此菌株具有較理想的遺傳穩(wěn)定性。4、通過復(fù)合誘變試驗,確定了UV+NTG復(fù)合誘變的條件:NTG的最佳濃度為0.6 mg/mL,15 w紫外燈,距離30 cm,照射處理時間120 s。該條件下的誘變效應(yīng)為:孢子致死率80.8%,正突變率8.76%。誘變后,獲得一株高產(chǎn)纖維素酶菌株-綠色木霉ZN4(Trichoderma viride.ZN4)。搖瓶發(fā)酵后,CMCase為1678.78U/mL,FPA為1031.56 U/mL,其產(chǎn)酶能力較出發(fā)菌株提高了19.1%和12.3%。經(jīng)傳代驗證,該菌株具有較理想的遺傳穩(wěn)定性和產(chǎn)酶潛力。5、利用出發(fā)菌株綠色木霉L4C和誘變菌株ZN4發(fā)酵處理秸桿3d、4d、5d后,結(jié)果表明:誘變菌株的效果優(yōu)于出發(fā)菌株,干物質(zhì)中粗蛋白的提高率在5 d時達81.69%,粗纖維的降解率在5 d時達39.98%。
[Abstract]:The optimum conditions for producing enzyme were studied by means of single factor test and orthogonal test . The optimum conditions were as follows : 1 . The optimum conditions for producing enzyme were obtained by means of orthogonal experiments . The optimum conditions for producing enzyme were as follows : 1 . The optimum conditions were as follows : 1 . Under optimum enzyme production conditions , CMCase was 921.56U / mL , and the initial pH was 5.0 . Under the condition of optimum enzyme production , the optimum conditions were as follows : 1 . The optimum conditions for producing enzyme were as follows : 1 . Under the optimum enzyme production condition , the optimum conditions were as follows : 1 . The optimum conditions for producing enzyme were as follows : The lethal rate was 85.0 % . After the mutation , a high yield cellulase strain , Trichoderma viride . Z15 was obtained . After shake flask fermentation , CMCase was 1196 . 21 U / mL , FPA was 863.56U / mL , and its enzyme - producing ability increased by 1.30 and 1.16 times respectively . After fermentation , CMCase was 1410 . 02 U / mL , FPA was 992.67U / mL . The yield of NTG was improved by 18.5 % and 9.7 % . The optimum concentration of NTG was 0.6 mg / mL , 15 w UV lamp , 30 cm distance and 120 s irradiation treatment time . The mutagenic effect of NTG was 80.8 % , positive mutation rate 8.76 % . After mutagenesis , a high yield cellulase strain , Trichoderma viride . ZN4 was obtained . After fermentation of shaking flask , CMCase was 1678.78U / mL , FPA was 1031 . 56 U / mL , and its enzyme - producing ability was improved by 19.1 % and 12.3 % . The results showed that the strain had better genetic stability and enzyme - producing potential .

【學(xué)位授予單位】：甘肅農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：S816

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