黄绿木霉内切葡聚糖酶ⅱ基因的克隆与表达-生物学；微生物学专业论文.docxVIP

目 目 录 4．2．1基因表达系统的选择 ～53 4．2．2制约外源基因表达的因素 ．．54 4．2．3关于构建高效表达载体方面的讨论 54 4．2．4内切葡聚糖酶诱导表达的研究 一55 4．2．5内切葡聚糖酶酶学特性分析 ～55 4．2．6内切葡聚糖酶发酵条件的优化 55 5结论 56 致谢 ．57 参考文献 ．58 攻读硕士学位期间发表的学术论文 65 III 万方数据 CONTENTSCoNTENTS CONTENTS CoNTENTS Abstract in Chinese ·l Abstract in English ． ．． ．．．．． ． ．．．．．．．．．．．．．． ．．．．．．．．． ．．．．．．．．． ．．．．．．．．．．． ·III 1．Introduction l 1．1 Cellulose and cellulases ．1 1．1．1 Cellulose ．．1 1．1．2 Cellulases ．2 1．2 Source of cellulases ．．4 1．2．1 Ceilulases from fungi ．4 1．2．2 Cellulases from bacterium 4 1．2．3 Cellulases from Actinomycete 4 1．2．4 Cellulases from animals ．5 1．3 Research actuality ．5 1．4 Application ofcellulases in industry ．6 1．g．1 In textile industry ．6 1．4．2 In food industry ．6 1．4．3 In feed industry ．6 1．4．4 In bioenergy industry ．．7 1．4．5 In papermaking industry 7 1．5 Aims ofresearch 7 1．6 Technological route ．．8 2．Materials and Methods．．．．．．． ．．．．．．．．．．．．．．．．．．．．．．．．．．．．．．．．．．．． ．．．．． ．．．．．．．．．．．．．．．．．．．．．．．．．． ．．．．．．．．．．．．．．． ．．．．9 2．1 Experimental materials 9 2．1．1 Strains and plasmids 9 2．1．2 Reagents and enzymes ． ． ． ． ll 2．1．3 Reagents and formula 一ll 2．1．4 Experimental Instruments ． ． ． ． ． ．． ． ． ． ． ． ． ． ． ． ． ． ． ． ． ． ．19 2．2 Experimental methods ．．14 2．2．1 Cultivation ofTrichoderma atroviride ．14 2．2．2 Cloning ofendoglucanase gene 一14 2．2．3 Bioinformatics analysis ．．1 8 2．2．4 Construction of integration vector ．1 8 2．2．5 Digestion and connection ． ．．21 2．2．6 TransfoITIIation ofSaccharomyces cerevis／ae 23 2．2．7 Induced expression and analysis ofEndoglucanase 24 2．2．8 Induced expression and analysis ofrecombinant enzyme 25 V 万方数据 东北农业大学理字坝士学位论义2．2．9 东北农业大学理字坝士学位论义 2．2．9 Enzyme properties and determination of kinetic parameters． ．．．．．．．．．．．．．．． ．．．．．． ．25 2．2．10 Recombinants optimization offermentation conditions ． ． ．．28 3．Results and analysis ． ． ． ． ．．． ．．．．．．．． ．．．．．．．．．．． ．．． ． ．．． ．．．．．．．．．．．． ．．．．．．． ． ． ．．．．．．．．．30 3．1 Cloning ofendoglucanase gene． ． ． 30 3．1．1 Extraction