Screening of the effective cellulose-degradable strain and its application in the production of cellulose bioethanol

Strains from the cellulose-containing environment were collected. Primary screening(by filter-paper Hutchison solid culture medium and sodium carboxymethylcellulose solid culture medium) and reelection(by filter-paper inorganic salt culture medium and sodium carboxymethylcellulose Congo red culture medium) indicated that five strains obtained were best suited for high performance cellulose degradation. Determination of sodium carboxymethylcellulose activity(CMCA) and filter paper activity(FPA) was accomplished for each of the five. The strongest of the five in CMCA and FPA was applied to the production of cellulose bioethanol by separate hydrolysis and fermentation(SHF) and simultaneous saccharification and fermentation(SSF) respectively.

Screening and Identification of Strains Converting Trans-anethole to Anisic Acid

[Objectives] This study was conducted to isolate and screen the bacteria that can convert trans-anethole to anisic acid from star anise and its environmental samples, and identify the bacteria. [Methods] According to the traditional microbial culture method, with trans-anethole as the sole carbon source, through enrichment culture and separation and purification, preliminary screening by thin layer chromatography and re-screening by high-performance liquid chromatography, strains that degraded trans-anethole to produce anisic acid were obtained, and 16 S rDNA sequencing and phylogenetic tree construction were performed for genetic analysis. [Results] Eleven strains that degraded trans-anethole to produce anisic acid were obtained, among which strain NT2 that produced anisic acid with a relatively high efficiency was initially identified as Pseudomonas sp. The strain’s trans-anethole degradation rate was 45.41%, and the molar production rate and cumulative concentration of anisic acid were 21.80% and 1.96 g/L, respectively. [Conclusions] Strain NT2 has a strong ability to degrade trans-anethole to produce anisic acid, and can enrich strain resources for degradation of trans-anethole to anisic acid through microbial conversion.

Primary Screening of 10-Hydroxy-2-Decenoic Acid Productive Strains

In this paper, eleven strains, which were screened strictly from raw royal jelly, soil andhoneycomb etc. by meansof dilution plate and spread plate methods, were cultured at 28℃ for 60 h with shaking. To determine whether they could yield10-Hydroxy-2-decenoic acid during fermentation, gas chromatography and gas chromatography-mass spectrometry methodswere used. The results showed that the strains BH002 and BH004, were both identified as Crytococcaceae, where BH002was primarily classified into Candida for possessing the abilities. The 10-HDA productivity of Candida BH002 and that ofBH004 were 0.327% and 0.2648% respectively.

Screening and degradation characteristics of a tylosin-degrading strain

Antibiotics residues have been accumulating in the environment day by day due to overuse of antibiotics.Recalcitrant antibiotic residues,such as tylosin(TYL),can cause serious environmental problems,which makes it important to eliminate TYL from the environment.It is important to eliminate TYL from the environment.In this study,a strain was isolated and purified from fermentation by-product that came from a TYL production factory.The TYL degrading strain was identified by its morphology,physiological and biochemical reactions and sequencing the PCR-amplified fragments of its 16 S r DNAcoding genes.The temperature,shaking speed,initial TYL concentration,p H and inoculum sizes were investigated under simulated conditions by using single factor tests.The results showed that TYL2,a high efficient strain was isolated and was identified as Brevibacillus borstelensis.The degradation rate of TYL by this strain could reach to 75%with an initial concentration of 25 mg L^-1 within 7 days under conditions of 7%B.borstelensis(v/v,2×108 CFU m L^-1)at p H 7.0 and at 35°C.It is interesting that this strain has a very strong ability to degrade the TYL in natural sewage with the degradation rate of 65%within 7 days.This result could be helpful for the degradation of TYL and provide guidance for the degradation of other antibiotics.

Screening influence on the Stark effect of impurity states in strained wurtzite GaN/Al_xGa_(1-x)N heterojunctions under pressure

The screening effect of the random-phase-approximation on the states of shallow donor impurities in free strained wurtzite GaN/Al x Ga 1 x N heterojunctions under hydrostatic pressure and an external electric field is investigated by using a variational method and a simplified coherent potential approximation.The variations of Stark energy shift with electric field,impurity position,Al component and areal electron density are discussed.Our results show that the screening dramatically reduces both the blue and red shifts as well as the binding energies of impurity states.For a given impurity position,the change in binding energy is more sensitive to the increase in hydrostatic pressure in the presence of the screening effect than that in the absence of the screening effect.The weakening of the blue and red shifts,induced by the screening effect,strengthens gradually with the increase of electric field.Furthermore,the screening effect weakens the mixture crystal effect,thereby influencing the Stark effect.The screening effect strengthens the influence of energy band bending on binding energy due to the areal electron density.

Screening and Preservation of Douchi Fibrinolytic Enzyme Producing Strain

[Objective] To isolate and preserve a high-activity Douchi fibrinolytic enzyme producing strain from Douchi products.[Method] The Douchi fibrinolytic enzyme producing strain was screened on the selected medium prepared with self-made pork blood powder,the strain with the highest activity was screened out according to the size of hydrolysis cycle,and then preserved in LB medium.[Result] A Douchi fibrinolytic enzyme producing strain with high thrombolytic activity was successfully screened out from the Douchi produced in Hunan,Guangdong and Jiangxi Provinces.[Conclusion] The study lays foundation for the development of new-type thrombolytic drugs.

Screening of Strains Producing Alkaline Protease from Soil and Study on the Conditions for Enzyme Production

[Objective] The aim was to screen strains producing alkaline protease from soil and study the conditions for enzyme production.[Method] Eight strains producing alkaline protease were isolated from soil through plate isolation,and the ability of enzyme production was measured by filter paper and Folin-phenol method.The strain with the strongest ability of enzyme production was screened as a candidate strain,then the factors influencing the ability of enzyme production was studied,finally the conditions for enzyme production was optimized through orthogonal test.[Result] No.5 strain was screened as a candidate strain due to its strong ability of enzyme production(6.00 U/ml),which accounted for 134.1% of that of Bacillus licheniformis,and it was gram-positive bacterium belonging to Clostridium.Orthogonal test showed that the optimal condition for producing protease was an environment with pH=11,0.3% of sucrose and 0.8% of peptone in the fermentation medium,and inoculation amount was 105 cfu/ml.In addition,peptone had significant impact on the level of enzyme production.[Conclusion] The study could provide theoretical references for the screening of strains producing alkaline protease.

Stretchable Strain Sensors Fabricated by Screen Printing of Silver Paste on the Surface Modified Transparent Elastomeric Polyurethane Films

Strain sensors for human-motion detection must offer high stretchability, high sensitivity, fast response, and high recovery speed. In this study, we choose silver paste as a sensing material and use a screen printing method to fabricate the strain sensor based upon an electrical-resistance mechanism. After curing elastomeric polyurethane film with a thickness of 150 μm on PET film, the polyester resin mixed with blocked isocyanate curing agent was coated as a masking layer to reduce the film’s stickiness. The effect of the polyester masking layer upon the silver paste screen printing process was examined using a rolling-ball-tack test, TGA analysis of polyester resins, and cured silver-electrode films. The cost-effective strain sensor fabricated by using silver paste and screen printing processes on the stretchable-polyurethane-substrate film showed high sensitivity and fast response in a strain range of up to 100%.

高温大曲中功能红曲霉菌株的筛选与功能浅析

该研究采用可培养技术从呈粉红色的高温大曲中分离红曲霉(Monascus)菌株,结合多元统计分析方法研究其在纯培养麦曲、模拟白酒固态发酵及强化大曲中的代谢特征,进而筛选优良的功能红曲霉菌株,并通过形态学观察和分子生物学技术对筛选菌株进行鉴定。结果表明,共分离筛选得到3株优良的功能红曲霉菌株,编号分别为H14、H26和H30。其中,菌株H14具有提高基酒产率(7.5%vol)、改善糟醅挥发性风味物质含量(33.12 mg/kg)和轮廓的潜力;菌株H30改善了高温大曲的酯化活性;菌株H26具有产特征风味化合物(吡嗪类物质)的潜力,其强化高温大曲中2,3,5,6-四甲基吡嗪的含量达0.27 mg/kg。经鉴定,菌株H14、H26和H30均为佛罗里达红曲霉(Monascus floridanus)。

高产苯乳酸菌株选育及发酵豆粕工艺优化

该研究以泡菜为原料,以金黄色葡萄球菌(Staphylococcus aureus)与大肠杆菌(Escherichia coli)为指示菌,通过溶钙圈法及牛津杯法初筛,苯乳酸产量检测复筛,从泡菜中分离筛选具有抑菌性能的高产苯乳酸菌株,将其应用于发酵豆粕以评估其产苯乳酸性能,并对其进行形态学观察、生理生化实验及分子生物学鉴定。结果表明,共获得8株具有抑菌效果的菌株,其中,菌株R69苯乳酸产量最高,为1.25 mg/mL,添加苯丙酮酸时,该菌株固态发酵豆粕的苯乳酸产量为425.70 mg/kg,该菌株被鉴定为植物乳杆菌(Lactobacillus plantarum)。菌株R69固态发酵豆粕时,添加0.2%蛋白酶、3%糖蜜和0.15%苯丙酮酸可提高苯乳酸含量,苯乳酸含量达到624.49 mg/kg,且可有效抑制大肠杆菌和霉菌的生长。

复合菌群构建及其石油烃降解特性

为获得高效降解石油烃复合菌群,对实验室保存的菌株进行石油烃降解效果及原油4组分降解能力分析研究,筛选出了4株菌(A1、A3、Y3、Y4),对其进行ITS和16S rRNA分子鉴定,确认4株菌的分类信息,并优化高效降解菌株的复配方式。结果显示:A1属于不动杆菌属(Acinetobacter)、A3属于芽胞杆菌属(Bacillus)、Y3和Y4均为假丝酵母菌属(Candida);A1、A3、Y3、Y4对石油烃的降解率分别为42.27%、40.95%、62.02%、69.84%;其中Y4对烷烃和芳烃降解效果最佳,降解率分别为65.71%和23.69%,沥青质降解效果最好的是A1,胶质降解效果最好的是Y3,降解率分别为22.74%和21.22%,A3对各组分降解能力均衡;A1、A3、Y4复合的菌群降解效果最佳,其降解率高达83.11%,相较于单一菌株提高了1.19—2.03倍;A1、A3、Y4的最佳接种量比例为0.9∶0.9∶1.5,其降解率可达88.34%。该研究成功筛选出一种新的高效降解石油烃的菌群,对进一步研究和应用于石油污染治理具有重要指导意义。

西藏牦牛瘤胃源产纤维素酶菌株筛选鉴定及产酶条件探究

为提高纤维素的利用率,寻找到安全性能好、产酶效率高的菌株添加剂添加到饲料中,以提升动物对饲料的利用率,从牦牛瘤胃中分离高产纤维素酶菌株,进行16S rDNA的鉴定,并探究其在不同温度、pH、接种量和发酵阶段产内切型-β-葡聚糖酶、外切型-β-葡聚糖酶、β-葡萄糖苷酶的特性。结果表明:通过形态学鉴定以及16S rDNA基因序列测定,最终确定高产菌株M1为克雷伯氏菌(Klebsiella oxytoca)。M1产内切型-β-葡聚糖酶活性大于外切型-β-葡聚糖酶、β-葡萄糖苷酶;M1在35℃、pH为7、接种量为2%的条件下产内切型-β-葡聚糖酶的效率最高,在30℃、pH为6、接种量为2%的条件下产外切型-β-葡聚糖酶的效率最高,在30℃、pH为6、接种量为1%条件下产β-葡萄糖苷酶的效率最高;在M1生长过程中,先产生了β-葡萄糖苷酶,其次是外切型-β-葡聚糖酶、内切型-β-葡聚糖酶,并且外切酶活性远远大于内切酶和β-葡萄糖苷酶活性。

降解氨基甲酸乙酯酿酒酵母的筛选及其应用

以宁夏5个产区的酿酒葡萄为原料筛选可以降解氨基甲酸乙酯(Ethyl carbamate,EC)且具有优良酿酒特性的红葡萄酒和白葡萄酒酿酒酵母。初筛试验获得了10株酵母,复筛试验以EC作为唯一氮源筛选出6株EC降解能力较好的酵母菌株,最后通过产气试验和耐受性试验筛选出了2株发酵性能较好的菌株。经过18S rDNA分子生物学鉴定后确定这2个菌株为酿酒酵母。使用筛选出的2个菌株发酵红葡萄酒和白葡萄酒,并以商业酵母作为对照,测定其理化指标及EC含量;再通过GC-MS分析其香气物质并结合感官评价最终筛选出具有优良酿酒特性和降解EC功效的红葡萄酒酿酒酵母(Saccharomyces cerevisiae)NXSC2和白葡萄酒酿酒酵母(Saccharomyces cerevisiae)NXSC1。

两株代谢咖啡酸人肠道来源菌株的筛选、鉴定及其代谢过程初探

为了探索咖啡酸的体内代谢过程,本研究从人体肠道粪便中分离筛选出2株能代谢咖啡酸的菌株,通过细胞形态、16S rDNA序列和系统发育树分析了其菌株特征,并研究了两株菌株代谢咖啡酸过程中的细菌总数和pH变化,最后采用薄层层析法和高效液相色谱法对代谢产物进行确定,阐明咖啡酸代谢过程。结果表明,2株菌分别为腐生葡萄球菌(Staphylococcus xylosus)和奇异变形杆菌(Proteus mirabilis)。腐生葡萄球菌代谢咖啡酸含量从0.539 mg/mL在6 h后降至0.087 mg/mL,12 h后完全被代谢。而奇异变形杆菌,在6 h后咖啡酸含量从0.543 mg/mL降至0.352 mg/mL,24 h后完全被代谢。咖啡酸抑制了肠道菌的增殖,且腐生葡萄球菌代谢咖啡酸能力优于奇异变形杆菌,肠道菌代谢咖啡酸产物为间香豆酸。研究结果为进一步研究咖啡酸和肠道菌的相互作用提供依据。

固态发酵制备黄精多糖的工艺优化、理化特性及抗氧化活性

选取7种乳杆菌进行固态发酵,以黄精多糖得率为评价指标确定最佳发酵菌种。借助单因素试验和响应面曲面分析法对固态发酵黄精工艺进行优化,并分析固态发酵黄精多糖(solid state fermentation Polygonatum sibiricum polysaccharide,SF-PSP)与相同条件下未经发酵黄精多糖(P.sibiricum polysaccharide,PSP)的单糖组成、形态特征、热失重特性以及体外抗氧化活性。结果表明:副干酪乳杆菌ATCC 334为最优发酵菌种,SF-PSP的最佳制备工艺条件为:液料比0.98∶1(mL∶g)、接菌量4.80%、发酵时间51.30 h,此条件下SF-PSP得率为(11.14±0.89)%。SF-PSP主要由甘露糖、半乳糖醛酸、半乳糖和葡萄糖组成,与PSP相比,鼠李糖、半乳糖醛酸、半乳糖含量均提高了4倍。扫描电镜结果表明,与PSP相比,SF-PSP结构更加疏松且有更多致密的小孔。热重分析结果表明2种多糖失重温度节点不同,且SF-PSP保水能力优于PSP。体外抗氧化活性结果表明SF-PSP的DPPH自由基清除率、ABTS阳离子自由基清除率、亚铁离子螯合能力及总抗氧化性均显著高于PSP(P<0.05)。综上,采用SF-PSP可以显著提高黄精多糖的得率和抗氧化活性。

基于Fosmid文库筛选山羊瘤胃微生物源蛋白酶基因及其表达验证

【目的】利用Fosmid文库功能筛选法,获得山羊瘤胃微生物源蛋白酶基因并进行原核表达。【方法】利用功能底物筛选法从山羊瘤胃微生物源Fosmid文库中筛选具有蛋白酶活性的克隆子,对其进行Illumina二代测序,对预测到的基因进行功能注释,根据基因丰度结果确定后续研究的功能基因。利用大肠杆菌BL21(DE3)对功能基因进行诱导表达,对表达产物进行酶活性测定。通过CRISPR/Cas9基因编辑技术,将密码子优化后的功能基因分别敲入枯草芽孢杆菌C6基因组ctc位点并进行表达验证。【结果】从Fosmid文库1700个克隆子中筛选到1个具有蛋白酶活性的阳性克隆Pro4-C5。通过二代测序技术,鉴定出3个潜在的蛋白酶基因gene0833(内肽酶,EC编号:EC3.4.21.53)、gene0196(金属内肽酶,EC编号:EC3.4.24)和gene0585(羧肽酶,EC编号:EC3.4.17.14)以及1个L-天冬酰胺酶基因(gene0683,EC编号:EC3.5.1.1)。以pET-28a(+)为表达载体,通过大肠杆菌BL21(DE3)原核诱导表达发现,gene0585和gene0196未表达;gene0833表达的蛋白分子质量为87 ku,蛋白酶活性为10.45 U/mL;gene0683表达的蛋白分子质量为37 ku,L-天冬酰胺酶活性为88.52 U/mL,同时发现该蛋白也具有蛋白酶活性,酶活性为5.25 U/mL。将密码子优化后的gene0683和gene0833定点敲入枯草芽孢杆菌C6基因组中表达发现,gene0833未表达,gene0683表达的蛋白酶活性为109.72 U/mL,相比原始菌株(100.97 U/mL)显著提高了8.67%(P<0.01);L-天冬酰胺酶活性为31.63 U/mL,相比原始菌株(22.79 U/mL)显著提高了30.06%(P<0.01)。【结论】从山羊瘤胃微生物源Fosmid文库中筛选和鉴定出了2个具有蛋白酶活性的功能基因(gene0833和gene0683),均来源于微小杆菌属(Exiguobacterium),其中gene0683在大肠杆菌和枯草芽孢杆菌中的表达产物具有较高的蛋白酶和L-天冬酰胺酶酶活性。

番茄青枯病拮抗菌的筛选及防治效果

为了筛选出具有抑制青枯雷尔氏菌作用的拮抗菌,采用形态指标、生理生化特征以及16S rRNA序列分析等方法对菌株进行了研究,并且通过阴性对照(CK)、番茄青枯病菌阳性胁迫处理(RB)和拮抗菌K+番茄青枯病菌处理(KRB)3个盆栽番茄土壤样品进行了防卫反应基因的表达研究.结果表明,拮抗菌K5菌落呈乳白色且表面光滑,经革兰氏染色为阳性,初步确定为芽孢杆菌,通过分子鉴定后确定为贝莱斯芽孢杆菌;番茄青枯病的盆栽防治效果达到23.20%,K5菌株在接种48 h后能诱导番茄根部抗病基因表达,CTR、ETR基因的表达量提高,表明K5通过植物抗病相关基因的表达,能诱导植物产生抗病性,并能有效地抑制青枯病菌的繁殖,从而达到防治青枯病的作用.

乙醇降解乳酸菌的筛选、鉴定及解酒防醉功效评价

该研究采用传统培养分离法从传统发酵食品奶疙瘩样品中分离乳酸菌,通过体外降解乙醇试验筛选高效降解乙醇功能的菌株,通过形态学观察、生理生化实验及分子生物学技术对其进行菌种鉴定,并对其进行安全性评价、耐逆性试验,以及体内小鼠防醉解酒功效试验。结果表明,筛选鉴定出一株乙醇降解率高(13.38%)、乙醇耐受能力强(耐受体积分数12%乙醇)的乳酸片球菌(Pediococcus acidilactici)RH2712,其对氨苄西林、青霉素、亚胺培南、氯霉素敏感,溶血性安全;对酸和胆盐耐受力强,在pH 3.0的条件下活菌对数值几乎不变,延迟时间为2 h,在1.5%胆盐含量下培养4 h后活菌对数值达到8.72,延迟时间仅为0.5 h,在人工模拟胃液条件下处理3 h后活菌对数值仍能达到7.14。乳酸片球菌RH2712在小鼠试验中表现出明显的防醉解酒的功效,且活菌数为1×10^(8)~1×10^(9)CFU/m L时解酒功能强于美他多辛。

耐盐碱性蛋白酶菌株LK-3的筛选及酶学性质

为了寻找产碱性蛋白酶活性高且稳定性良好的野生菌株,作者从上海临港区域的东海海水中筛选出一株高产稳定且耐盐的产碱性蛋白酶菌株。对该菌株的形态学特性、生理特性以及16S rDNA基因序列进行测序和分析,确定该菌株为同温层芽孢杆菌(Bacillus stratosphericus),并命名为B. stratosphericus LK-3。酶学性质研究结果显示,该酶的最适作用条件为温度35℃、p H 8.5、最适接种体积分数7%,该酶在上述条件下发酵72 h的酶活力为(581.74±0.81) U/mL。此外该酶具有较好的耐盐性,即使在40 g/dL的高饱和盐质量浓度下仍然保持22.03%的相对原始酶活力。蛋白酶的耐盐性是一个有价值的特性,可将其纯化后作为洗涤剂的生物添加剂应用于工业生产。

小站稻秸秆降解复合菌系的筛选与产酶条件优化

为寻找一种能够科学高效地将小站稻秸秆用于秸秆还田的方法,并将其作为一种储备技术,本研究通过测定已有6组秸秆降解复合菌系的羧甲基纤维素酶活力和滤纸酶活力,筛选出2组复合菌系,并通过正交试验进行产酶条件的优化,以探索解决小站稻秸秆清洁生产技术的应用问题,保障小站稻的未来绿色可持续发展,为小站稻秸秆高效处理的研究提供理论基础和技术支持。本研究结果表明:复合菌系B和复合菌系F的综合酶活力均高于其他复合菌系;复合菌系B羧甲基纤维素酶最佳发酵条件为25℃,pH 8.0,接种量2%,温度为酶活力主要影响因素;复合菌系B滤纸酶最佳发酵条件为20℃,pH 9.0,接种量2%,pH为酶活力主要影响因素;复合菌系F羧甲基纤维素酶最佳发酵条件为25℃,pH 8.0,接种量2%,温度为酶活力主要影响因素;复合菌系F滤纸酶最佳发酵条件为20℃,pH9.0,接种量2%,接种量为酶活力主要影响因素。