CN116200286B - 一株高效糖化纤维素的热纤梭菌及其应用 - Google Patents
一株高效糖化纤维素的热纤梭菌及其应用 Download PDFInfo
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Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
- C12N1/205—Bacterial isolates
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/02—Monosaccharides
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/12—Disaccharides
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
- C12R2001/00—Microorganisms ; Processes using microorganisms
- C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
- C12R2001/145—Clostridium
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
Abstract
一株高效糖化纤维素的热纤梭菌及其应用,涉及微生物领域,尤其涉及一株热纤梭菌及其应用。是要解决现有的热纤梭菌难以积累可溶性糖,糖化率较低的问题。该糖化纤维素的热纤梭菌为热纤梭菌FC811,保藏在中国微生物菌种保藏管理委员会普通微生物中心,保藏日期为2021年12月23日,保藏编号为CGMCC No:40026。本发明热纤梭菌FC811能够降解纤维素又能够积累可溶性糖,糖化率可接近60%。本发明的热纤梭菌用于降解纤维素并积累可溶性糖。
Description
技术领域
本发明涉及微生物领域,尤其涉及一株热纤梭菌及其应用。
背景技术
木质纤维素主要包括农业固体废物、花草树木以及一些经济作物等,是自然界中一类分布范围最广,数量巨大且能够再生的生物质。这些生物质如果处理不当不仅会造成资源的浪费还会严重污染环境。因此,合理转化和利用这些生物质资源,对于经济开发和建设环境友好型社会具有重要意义。然而,由于木质纤维素结构复杂,因此,如果能够找到一种高效利用木质纤维的菌株非常重要。
热纤梭菌是能够高效降解木质纤维素的一类厌氧细菌。然而,目前能够降解纤维素的热纤梭菌,尽管有较多秸秆糖化菌株研究,由于它们为了维持自身的生长,需要大量消耗糖,因葡萄效应的存在而极难在培养体系中积累可溶性糖,因此糖化率较低,现有的热纤梭菌糖化率最高能达到49%。因此,筛选出一株能够高效糖化纤维素的热纤梭菌,对于农业废弃物资源化利用具有重要意义。
发明内容
本发明是要解决现有的热纤梭菌难以积累可溶性糖,糖化率较低的问题,提供一株高效糖化纤维素的热纤梭菌及其应用。
本发明高效糖化纤维素的热纤梭菌为热纤梭菌(Clostridium thermocellum)FC811,保藏在中国微生物菌种保藏管理委员会普通微生物中心(CGMCC),保藏地址是北京市朝阳区北辰西路1号院3号,保藏日期为2021年12月23日,保藏编号为CGMCC No:40026。
本发明热纤梭菌(Clostridium thermocellum)FC811在微晶纤维素滚管培养基上能够形成透明的水解圈,在60℃培养箱培养一段时间后,水解圈明显变大,同时在水解圈中间可发现黄色菌落,表面光滑,不透明,直径约为0.5-2mm。
所述微晶纤维素滚管培养基配方为:氯化钠1.0g/L、氯化铵1.0g/L、磷酸氢二甲1.5g/L、磷酸二氢钾3.5g/L、氯化镁0.5g/L、氯化钾0.2g/L、酵母提取物2g/L、蛋白胨2g/L半胱氨酸0.6g/L、琼脂粉20g/L、微量元素5ml/L、维生素0.5ml/L、0.1%(w/v)刃天青0.2g/L、微晶纤维素5g/L、蒸馏水1000ml,pH值调至7,121℃灭菌15min。
本发明热纤梭菌FC811为革兰氏阳性菌,呈杆状,有芽孢,有鞭毛,能运动,单个生长。在厌氧条件下生长,最适生长温度为60℃。热纤梭菌FC811的运动性为阳性,葡萄糖试验为阳性,甘露糖试验为阳性,微晶纤维素试验为阳性;甘露醇试验为阳性,纤维二糖试验为阳性,乳糖试验为阴性,果糖试验为阳性,蔗糖试验为阳性,麦芽糖试验为阳性,木聚糖试验为阳性,木糖试验为阴性,淀粉试验为阴性,滤纸试验为阳性,明胶试验为阳性,硝酸盐还原试验为阴性,柠檬酸盐试验为阴性,羧甲基纤维素钠试验为阳性,荧光色素试验为阴性。
本发明热纤梭菌FC811通过16S rDNA序列比对分析,与热纤梭菌(Clostridium.thermocellum strain JN4)的相似性高达100%,与模式菌株热纤梭菌(R.thermocellum ATCC 27045)的相似性也为100%,结合菌体的形态学观察、生理生化分析,确定菌株FC811为热纤梭菌(Clostridium thermocellum),命名为Clostridiumthermocellum FC811。
本发明还提供热纤梭菌FC811在降解纤维素中的应用。
进一步的,所述纤维素为微晶纤维素。
本发明还提供热纤梭菌FC811在降解纤维素过程中积累糖中的应用。
进一步的,所述糖为可溶性糖。
本发明的有益效果:
本发明热纤梭菌FC811能够降解纤维素又能够积累可溶性糖。在以微晶纤维素为底物时,可发现血清瓶中有许多黄色亲和力物质(YAS)生成,这种物质是类胡萝卜素,能够促进微晶纤维素的降解。
本发明的热纤梭菌FC811不仅能够降解微晶纤维素,而且还能够积累大量的可溶性糖,经优化后其糖化率能够达到57.72%,明显高于以往报道的热纤梭菌。以往报道该属的菌株,对糖的利用率较高,因此,糖的积累量较低。然而,本发明筛选到的菌株之所以能够高效积累可溶性糖,是因为该菌株在生长过程中只需少量的糖类就能维持自身的生长繁殖,对糖的利用率较低,且菌株体内存在大量的纤维素酶,能够将纤维素转化成可溶性糖,因此,可溶性糖的积累量高。
本发明分离筛选出高效纤维素糖化菌株对秸秆材料资源化和能源化利用具有重要的战略意义,同时为其同步发酵产糖产氢的研究和进一步规模化生产应用提供参考。
附图说明
图1为热纤梭菌FC811在厌氧管表面的菌落形态;
图2为热纤梭菌FC811革兰氏染色菌体形态(×100);
图3为热纤梭菌FC811单菌扫描电镜照片;
图4为热纤梭菌FC811的系统发育树;
图5为温度对菌株FC811的生长和微晶纤维素糖化能力的影响;
图6为初始pH对菌株FC811的生长和微晶纤维素糖化能力的影响;
图7为底物浓度对菌株FC811的生长和微晶纤维素糖化能力的影响;
图8为酵母粉浓度对菌株FC811的生长和微晶纤维素糖化能力的影响;
图9为培养时间对菌株FC811的生长和微晶纤维素糖化能力的影响;
图10为菌株FC811降解微晶纤维素的糖化产物及高效液相色谱检测分析;其中曲线a表示葡萄糖,曲线b表示纤维二糖。
具体实施方式
本发明技术方案不局限于以下所列举具体实施方式,还包括各具体实施方式间的任意组合。
实施例1:本实施例热纤梭菌(Clostridium thermocellum)FC811,保藏在中国微生物菌种保藏管理委员会普通微生物中心(CGMCC),保藏地址是北京市朝阳区北辰西路1号院3号,保藏日期为2021年12月23日,保藏编号为CGMCC No:40026。
本实施例热纤梭菌FC811的获取方法如下:
所述热纤梭菌FC811分离自牛瘤胃残留物、森林腐木、原始森林30cm土壤、温泉底泥以及厌氧高温堆肥等环境样品的混合物中。具体分离方法为:将不同环境中的样品进行混匀,称取10g,放入盛有90mL无糖秸秆基础培养基和数粒玻璃珠的厌氧发酵瓶中,于60℃、150rpm水浴振荡富集一周。取富集悬液5mL接种于45mL微晶纤维素培养基中,培养1-2周后,再于无糖秸秆基础培养基中连续接种3-5次,得到产糖性能较为稳定的复合菌系FS-1。将富集筛选得到的复合菌系,接种于微晶纤维素培养基中,培养一周后,吸取悬液1mL进行10-1~10-5浓度梯度稀释,然后分别吸取10-3、10-4、10-5三个稀释度各1mL注入分离纯化培养基中,于60℃培养箱中静态培养一周后,可在分离纯化培养基表面发现多个大小不一的水解圈,在其中央可观察到菌落,分别将透明圈较大区域,长势良好的菌落用接菌针进行剥离,将其溶解到新的微晶纤维素培养基中,于60℃培养一周。重复以上步骤若干次,直到观察到厌氧滚管内微晶纤维素表面细菌形态相同,结合显微镜和扫描电镜,最后成功筛选分离到高效纤维素产糖菌株,即为本实施方式的热纤梭菌FC811。
所述无糖秸秆基础培养基的配方为:氯化钠1.0g/L、氯化铵1.0g/L、磷酸氢二甲1.5g/L、磷酸二氢钾3.5g/L、氯化镁0.5g/L、氯化钾0.2g/L、酵母提取物2g/L、蛋白胨2g/L、半胱氨酸0.6g/L、微量元素5ml/L、维生素0.5ml/L、0.1%(w/v)刃天青0.2g/L、秸秆5g/L、蒸馏水1000ml、pH值调至7。
所述微晶纤维素培养基的配方为:碳源为微晶纤维素,其他成分与无糖秸秆基础培养基相同,121℃灭菌15min。所述分离纯化培养基的配制过程为:向1L基础培养基中添加20g琼脂,配制成固体滚管培养基,往厌氧管中注射6mL融化后的固体培养基,经过121℃灭菌后,将其冷却到50~60℃时,放入装有冰水混合物的平底容器或水盆中,迅速的来回滚动,大约1~2min完全凝固,制成厌氧固体滚管;其配方与微晶纤维素培养基相同。
所述微量元素的配方为:氯化亚铁1.5g/L、氯化锌70.0mg/L、硼酸6.0mg/L、四水氯化锰0.1g/L、二水氯化铜2.0mg/L、六水氯化钴0.19g/L、六水氯化镍24.0mg/L、一水钼酸钠36.0mg/L、钨酸钠15.0mg/L、五水硒酸钠15.0mg/L、蒸馏水1000ml。所述维生素的配方为:硫辛酸50.0mg/L、生物素20.0mg/L、烟酸0.35g/L、盐酸硫胺素5.0mg/L、对氨基苯甲酸50.0mg/L、叶酸20.0mg/L、泛酸钙50.0mg/L、维生素B12 1.0mg/L、盐酸吡哆醇100.0mg/L、蒸馏水1000mg/L,pH值调至6.8-7.0。
实施例2:热纤梭菌FC811的理化特征和分子鉴定
(1)热纤梭菌FC811的形态特征:
热纤梭菌FC811在微晶纤维素滚管培养基上能够形成透明的水解圈,在60℃培养箱培养一段时间后,水解圈明显变大,同时在水解圈中间可发现黄色菌落,表面光滑,不透明,直径约为0.5-2mm。如图1所示。
所述微晶纤维素滚管培养基配方为:氯化钠1.0g/L、氯化铵1.0g/L、磷酸氢二甲1.5g/L、磷酸二氢钾3.5g/L、氯化镁0.5g/L、氯化钾0.2g/L、酵母提取物2g/L、蛋白胨2g/L半胱氨酸0.6g/L、琼脂粉20g/L、微量元素5ml/L、维生素0.5ml/L、0.1%(w/v)刃天青0.2g/L、微晶纤维素5g/L、蒸馏水1000ml,pH值调至7,121℃灭菌15min。
(2)热纤梭菌FC811的生理生化特征:
热纤梭菌FC811为革兰氏阳性菌,呈杆状,有芽孢,有鞭毛,能运动,单个生长,如图2和图3所示。在厌氧条件下生长,最适生长温度为60℃。热纤梭菌FC811的运动性为阳性,葡萄糖试验为阳性,甘露糖试验为阳性,微晶纤维素试验为阳性;甘露醇试验为阳性,纤维二糖试验为阳性,乳糖试验为阴性,果糖试验为阳性,蔗糖试验为阳性,麦芽糖试验为阳性,木聚糖试验为阳性,木糖试验为阴性,淀粉试验为阴性,滤纸试验为阳性,明胶试验为阳性,硝酸盐还原试验为阴性,柠檬酸盐试验为阴性,羧甲基纤维素钠试验为阳性,荧光色素试验为阴性。
表1热纤梭菌FC811生理生化特征
(3)热纤梭菌FC811的分子鉴定结果:
将FC811菌株接种注射到新配置的液态发酵培养基中,于60℃,150r/min摇床培养7d。所述液态发酵培养基的配方为氯化钠1.0g/L、氯化铵1.0g/L、磷酸氢二甲1.5g/L、磷酸二氢钾3.5g/L、氯化镁0.5g/L、氯化钾0.2g/L、酵母提取物2g/L、蛋白胨2g/L、半胱氨酸0.6g/L、微量元素5ml/L、维生素0.5ml/L、0.1%(w/v)刃天青0.2g/L、微晶纤维素5g/L、蒸馏水1000ml、pH值调至7,121℃灭菌15min。将厌氧菌株培养至对数期,取10mL菌液,500rpm离心2min后,弃去发酵后的微晶纤维素,取上清悬液,5000rpm离心3min,收集菌体,采用试剂盒提取基因组DNA,然后以基因组DNA为模板,采用通用引物27-F:5’-AGAGTTTGATCCTGGCTCAG-3’和1492-R:5’-TACGGCTTACCTTGT TACGACTT-3’进行16S rDNA的PCR扩增。扩增产物经纯化分离后,采用胶回收试剂盒回收,交由上海生工生物股份有限公司测序,经测序结果验证,FC811菌株16S rDNA序列长度为1464bp,如序列表中SEQ ID NO:1所示。对其基因序列与GenBank数据库中的序列进行相似性检索分析,获得与菌株FC811的16S rDNA同源性较高的序列,构建系统发育树,如图4所示。
FC811菌株16S rDNA序列如下:
GCTCAGGACGAACGCTGGCGGCGTGCCTAACACATGCAAGTCGAGCGGGGAT
ATACGGAAGGTTTACCGGAAGTATATCCTAGCGGCGGACGGGTGAGTAACGCGTGGGTAACCTACCTCATACAGGGGGATAACACAGGGAAACCTGTGCTAATACCGCATAACATAACGGGGCGGCATCGTCCTGTTATCAAAGGAGAAATCCGGTATGAGATGGGCCCGCGTCCGATTAGCTGGTTGGTGAGGTAACGGCTCACCAAGGCGACGATCGGTAGCCGAACTGAGAGGTTGGTCGGCCACATTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTGGGGAATATTGCGCAATGGGGGAAACCCTGACGCAGCAACGCCGCGTGAAGGAAGAAGGCCTTCGGGTTGTAAACTTCTTTGATTGGGGACGAAGGAAGTGACGGTACCCAAAGAACAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCGAGCGTTGTCCGGAATTACTGGGTGTAAAGGGCGCGTAGGCGGGGATGCAAGTCAGATGTGAAATTCCGGGGCTTAACCCCGGCGCTGCATCTGAAACTGTATCTCTTGAGTGCTGGAGAGGAAAGCGGAATTCCTAGTGTAGCGGTGAAATGCGTAGATATTAGGAGGAACACCAGTGGCGAAGGCGGCTTTCTGGACAGTAACTGACGCTGAGGCGCGAAAGCGTGGGGAGCAAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGGATACTAGGTGTAGGAGGTATCGACCCCTTCTGTGCCGGAGTTAACACAATAAGTATCCCACCTGGGGAGTACGGCCGCAAGGTTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCAGTGGAGTATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGGCTTGACATCCCTCTGACAGCTCTAGAGATAGGGCTTCCCTTCGGGGCAGAGGAGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGTCGTTAGTTGCCAGCACGTTAAGGTGGGCACTCTAGCGAGACTGCCGGCGACAAGTCGGAGGAAGGTGGGGACGACGTCAAATCATCATGCCCCTTATGTCCTGGGCTACACACGTACTACAATGGCTGCTACAAAGGGAAGCGATACCGCGAGGTGGAGCAAATCCCCAAAAGCAGTCCCAGTTCGGATTGCAGGCTGAAACTCGCCTGCATGAAGTCGGAATTGCTAGTAATGGCAGGTCAGCATACTGCCGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCATGAGAGTCTGCAACACCCGAAGTCAGTAGTCTAACCGCAAGGAGGGCGCTGCCGAAGGTGGGGCAGATGATTGGGGTGAAGTCGTA
菌株FC811与热纤梭菌(Clostridium.thermocellum strain JN4)的相似性高达100%,与模式菌株热纤梭菌(R.thermocellum ATCC 27045)的相似性也为100%,结合菌株FC811的形态学观察、生理生化分析,确定菌株FC811为热纤梭菌(Clostridiumthermocellum),命名为Clostridium thermocellum FC811。
实施例3:
(1)菌株FC811降解微晶纤维素的糖化产物及高效液相色谱检测分析
于100ml的厌氧瓶中加入45mL发酵培养基,微晶纤维素浓度为5g/L,以10%的接种量将菌株FC811的菌液注射到发酵培养基中,pH 7.0,在60℃,130rpm水浴恒温振荡器培养,培养时间为1d、2d、3d、4d、5d、6d、7d、8d,每个培养时间设置3个平行,培养后取样检测,采用高效液相色谱检测分析,检测溶液中的糖化产物及积累量,其结果如图10所示。
由图10可知,菌株FC811降解微晶纤维素的主要产物为葡萄糖和纤维二糖,随着发酵时间的增加,葡萄糖的积累量呈现出先上升后趋于稳定的变化趋势,而纤维二糖呈现出先上升后下降的变化趋势。当培养时间为8d,葡萄糖的积累量为540.4mg/g,纤维二糖的积累量为49.3mg/g,其总糖产量为589.7mg/g。因此,菌株FC811是一株高效纤维素产糖菌株。
(2)温度对菌株FC811纤维素糖化效果的影响
在100mL的厌氧瓶中,加入45mL含5g/L微晶纤维素的发酵培养基,经灭菌后,将纤维素糖化菌FC811以10%的接种量(v/v)注射到厌氧瓶中,于45℃、50℃、55℃、60℃、65℃、70℃、75℃条件下,pH 7.0,130rpm水浴恒温振荡器培养,每个温度设置3个平行,培养6d后取样检测,采用DNS法和紫外可见分光光度计测定波长540nm处的吸光度,计算菌株FC811的生物量和溶液中可溶性糖的积累量,其结果如图5所示,其中表示糖产量,/>表示糖化率,折线表示OD600。
(3)初始pH对菌株FC811纤维素糖化效果的影响
在100mL的厌氧瓶中,加入45mL发酵培养基,微晶纤维素浓度为5g/L,用1mol/LHCl和NaOH将pH值调为5.0、5.5、6.0、6.5、7.0、7.5、8.0、8.5、9.0,将纤维素糖化菌FC811以10%的接种量(v/v)注射到发酵培养基中,在60℃,130rpm水浴恒温振荡器培养,每个pH设置3个平行,培养6d后取样检测,采用DNS法和紫外可见分光光度计测定波长540nm处的吸光度,计算菌株FC811的生物量和溶液中可溶性糖的积累量,其结果如图6所示,其中表示糖产量,/>表示糖化率,折线表示OD600。
(4)底物浓度对菌株FC811纤维素糖化效果的影响
于100ml的厌氧瓶中,加入45mL发酵培养基,将发酵培养基中微晶纤维素的浓度分别调节为1.0、2.0、3.0、4.0、5.0、6.0、7.0g/L,纤维素糖化菌FC811接种量为10%,pH 7.0,在60℃,130rpm水浴恒温振荡器培养,每个浓度设置3个平行,培养6d后取样检测,采用DNS法和紫外可见分光光度计测定波长540nm处的吸光度,计算菌株FC811的生物量和溶液中可溶性糖的积累量,其结果如图7所示,其中表示糖产量,/>表示糖化率,折线表示OD600。
(5)酵母粉含量对菌FC811纤维素糖化效果的影响
于100mL的厌氧瓶中,加入45mL含5g/L微晶纤维素的发酵培养基,调节发酵培养基中酵母粉的浓度分别为0.0、0.5、1.0、1.5、2.0、2.5、3.0、3.5g/L,pH为7,温度为60℃,菌株FC811接种量为10%,培养时间为6d,130rpm水浴恒温振荡器培养,每个浓度设置3个平行,采用DNS法和紫外可见分光光度计测定波长540nm处的吸光度,计算菌株FC811的生物量和溶液中可溶性糖的积累量,其结果如图8所示,其中表示糖产量,/>表示糖化率,折线表示OD600。
(6)培养时间对菌株FC811纤维素糖化效果的影响
于100ml的厌氧瓶中加入45mL发酵培养基,微晶纤维素浓度为5g/L,以10%的接种量将纤维素糖化菌FC811的菌液注射到发酵培养基中,pH 7.0,在60℃,130rpm水浴恒温振荡器培养,培养时间为1d、2d、3d、4d、5d、6d、7d、8d,每个培养时间设置3个平行,培养后取样检测,采用DNS法和紫外可见分光光度计测定波长540nm处的吸光度,计算菌株FC811的生物量和溶液中可溶性糖的积累量,其结果如图9所示,其中表示糖产量,/>表示糖化率,折线表示OD600。
通过单因素实验得到,当菌株培养温度为60℃、pH为7.0、底物浓度为5.0g/L、酵母粉的浓度为2.0g/L时,热纤梭菌FC811具有最佳的糖化效果,可溶性糖的积累量可达0.57g/g微晶纤维素,按照以下公式计算糖化率,糖化率为57.72%。
以上所述的发酵培养基配方为:氯化钠1.0g/L、氯化铵1.0g/L、磷酸氢二甲1.5g/L、磷酸二氢钾3.5g/L、氯化镁0.5g/L、氯化钾0.2g/L、酵母粉2g/L、蛋白胨2g/L、半胱氨酸0.6g/L、微量元素5ml/L、维生素0.5ml/L、0.1%(w/v)刃天青0.2g/L、微晶纤维素5g/L、蒸馏水1000mL、pH值调至7。由于是单因素实验,其配方里的温度、pH、底物浓度、酵母粉以及培养时间等因素有所变化,其他成分保持不变。
Claims (5)
1.一株糖化纤维素的热纤梭菌,其特征在于所述糖化纤维素的热纤梭菌为热纤梭菌(Clostridium thermocellum)FC811,保藏在中国微生物菌种保藏管理委员会普通微生物中心,保藏地址是北京市朝阳区北辰西路1号院3号,保藏日期为2021年12月23日,保藏编号为CGMCC No:40026。
2.如权利要求1所述的热纤梭菌FC811在降解纤维素中的应用。
3.根据权利要求2所述的应用,其特征在于所述纤维素为微晶纤维素。
4.如权利要求1所述的热纤梭菌FC811在降解纤维素过程中积累糖的应用。
5.根据权利要求4所述的应用,其特征在于所述糖为可溶性糖。
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---|
刘世岳等.颜飞-基于全菌催化剂的木质纤维素整合生物糖化.《中国生物工程学会第十三届学术年会暨2019年全国生物技术大会论文集》.2019,第1-7页. * |
刘艳.嗜热厌氧梭菌降解纤维素及产氢特性研究.《中国博士学位论文全文数据库 基础科学辑》.2009,(第第05期期),A006-42. * |
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