Automated and integrated ultrahigh throughput screening for industrial strain enabled by acoustic-droplet-ejection mass spectrometry

Synthetic biology provides unprecedented opportunities to tackle critical issues including climate change and sustainable development by constructing microbial cell factories to produce industrially valuable biochemicals,biofuels,and biomaterials using renewable biomass resources[1],where strain screening for specific metabolic traits is a critical step.Rapid,accurate,and simultaneous quantification of multiple metabolites is critical for multi-perspective strain performance evaluation and strain screening,and a generalized method or platform will also reduce method development time to speed up the screening process.

Isolation,Screening and Identification of Cellulose Decomposing Strains in Straw-amended Soil

[Objective]The aim was to isolate cellulose decomposing strains with high activity from straw-amended soil.[Method]Screening of CMC solid culture and shaking culture rescreening were carried out to obtain cellulose decomposing strains with high activity from the corn straw amended soil,which were analyzed by 16rDNA sequence analysis.[Result]A bacterium and a fungus with higher endonucleases activity were obtained through preliminary screening and secondary screening,a fungus with higher filter paper enzyme activity,and a bacterium（NO.5 strain） with higher of both filter paper enzyme activity and CMC enzyme activity were obtained.The result of 16S rDNA sequence analysis showed that the similarity was 100% between NO.5 strain and Bacillus subtilis.[Conclusion]NO.5 strain was identified as Bacillus subtilis.

Isolating and Screening of Effective Decomposing Strain of Silkworm Chrysalis Protein and Study on Parts of Its Enzymatic Properties

[Objective] The research aimed to obtain an effectively-decomposing strain of silkworm chrysalis protein and discuss its enzymatic properties.[Method] The effectively-decomposing bacteria of protein was isolated from the decayed silkworm chrysalis by using dilution plate and its enzymatic properties were tested after primary screening and second screening.The enzyme activity was determined and the intermediate and small molecule protein content in silkworm chrysalis was measured after solid-state fermentati...

Screening and Molecular Identification of Strain that Produced Inulinase in Qinghai

The 46 strains that produced inulase were screened from rhizosphere soil where Jerusalem artichoke was planted in Qinghai.One strain with high inulinase productivity was obtained through further screening and the enzyme activity was6.67 U/ml.This inulinase was exonuclease.Through determination and analysis of the r DNA-ITS sequence,and analysis of morphology,the fungus was identified as Actinomucor elegans.

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/AlxGa1-xN 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, A1 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.

Electric-magnetic-force characteristics of rare earth barium copper oxide superconductor high-field coils based on screening effect and strain sensitivity

Rare earth barium copper oxide(REBCO)is the most researched and commercialized second-generation high-temperature superconducting material.Due to the anisotropic structure,strong deformation sensitivity,and central field errors caused by screening current effects,it is still a challenge for commercialization applications.In this study,the transversely isotropic constitutive relationship is selected as the mechanical model based on the structural characteristics of REBCO tapes,and suitable microelements are selected to equate the elastic constants using their average stress-strain relationships.Then,a two-dimensional axisymmetric model for coils wound by single-layer tapes is constructed to analyze the dependence of the electric-magnetic-force distribution in the tape on the strain.Finally,the anisotropic approximation of the homogenized bulk method is used to equate large-turn high-field coils,and the electric-magnetic-force distribution characteristics of the coils with/without screening effects and mechanical strain conditions are investigated,respectively.The results reveal that the mechanical strain has a weakening effect on the electromagnetic field distribution of superconducting tapes,but causes a significant enhancement in the force field distribution.In the presence of 0.5% mechanical strain,the maximum weakening of the peak value of the current density and the critical current density inside the high-field coil can reach about 8% and 13%,respectively,with a nearly 5 times increase in the peak stress.The screening current makes the current field distribution inside the coil improve by about 10 times.The screening current induced magnetic field can reach up to 0.8 T,making the relative error of the high-field coil center up to 7.8%.

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 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 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.

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 flbrinolytic 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. [ Rebait] A Douchi fibrinolytic enzyme producing strain with high thrombolytic activity was successfully screened out from the Douchi produced in Hunan, Guangdong and Jiangxi Prov- inces. [ Ceadusioe] The study lays foundation for the development of new-type thrombolytic drugs.

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.

Binding Energies of Screened Excitons in a Strained(111)-Oriented Zinc-Blende GaN/AlGaN Quantum Well Under Hydrostatic Pressure

We investigate the binding energies of excitons in a strained （111）-oriented zinc-blende GaN/Al0.3 Ga0.7 N quantum well screened by the electron-hole （e-h） gas under hydrostatic pressure by combining a variational method and a selfconsistent procedure. A built-in electric field produced by the strain-induced piezoelectric polarization is considered in our calculations. The result indicates that the binding energies of excitons increase nearly linearly with pressure,even though the modification of strain with hydrostatic pressure is considered, and the influence of pressure is more apparent under higher e-h densities. It is also found that as the density of an e-h gas increases,the binding energies first increase slowly to a maximum and then decrease rapidly when the e-h density is larger than about 1 ×10^11 cm^-2. The excitonic binding energies increase obviously as the barrier thickness decreases due to the decrease of the built-in electric field.

Study on Rational Breeding of High-Yield Avermectin Industrial Strain

[Objective] Strain Biok Av-023 used as the control was employed on screening of high-avermectin yield mutants by rational screening.[Method] With Biok Av-023 as the original strain,the positive mutation strain was firstly screened by routine UV mutagenesis,and then the high-yield avermectin producing strain was selected by the breeding way inferred by L-Ile induction.[Result] UV mutation and L-Ile directional screening had showed that the best L-Ile screening concentration was 0.5%,and the high-yield mutation strain AV60s-32 after re-screening reached the highest titer of 4520 IU/ml,which increased by 23.4% compared with the original strain.[Conclusion] The production of avermectin can be effectively enhanced by the combined way of UV mutation and L-Ile rational breeding.

Screening and Identification of A Probiotic from the Intestinal Tract of Mystus macropterus

A total of 25 strains were isolated from the intestinal tract of Mystus macropterus. The inhibitory effects of extracellular enzyme activity to common aquat- ic pathogenic bacteria were screened. A probiotic DQHY -7 strain was isolated, and its safety was detected. DQHY -7 strain was preliminarily identified by mor- phological observation, and physiological and biochemical test. Research results showed that DQHY -7 strain had relatively strong production capacities of amylase, cellulase and protease, and could restrict the growth of Escherichia coli, Aeromonas hydrophila and Pseudomonas fluorescens. Results of acute toxicity test showed that 1. 0 × 108 CFU/mL DQHY -7 bacterial suspension showed no pathogenicity to M. macropterus. According to the colony growth pattern and physiological-bio- chemical characteristics of DQHY -7, the strain was identified to be Bacillus subtilis. Research results showed that DQHY -7 strain could be used as a new candi- date strain for feeding microecologicalagent.

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%.

Screening of Highly-efficient Triethylamine-degrading Bacteria

Triethylamine is used in the production process in many fields,and it will enter the environment and cause pollution in the process of use.As a pollutant,triethylamine will cause great harm to aquatic organisms and affect the development of aquaculture.It is very important to degrade triethylamine.Traditional degradation methods have the disadvantages of low efficiency and high cost.It is safer,more effective and cheaper to degrade triethylamine by microbial decomposition.In this study,a strain(Microbacterium actinomyceterae CC-VM-Y(T))which can degrade triethylamine efficiently was successfully screened,providing an effective and economical method for degrading triethylamine and protecting aquaculture environment.

Study on Isolation and Degradation Characteristics of an Aniline-degrading Strain

[Objective] The research aimed to screen out an efficient aniline-degrading strain and study its degradation characteristics. [Method] By domesticated enrichment culture, an efficient aniline-degrading strain named as DA-K was isolated from activated sludge sample collected from a chemical plant in Henan. DA-K could use aniline as the sole carbon and nitrogen sources. The strain was carried out physiological and biochemi- cal identification, and its biological degradation characteristics were studied. [ Result] DA-K strain was G - and red-shaped, and its colony color was off-white. It was initially determined as Acinetobacter sp. By measuring, the optimum growth temperature and pH for DA-K were respectively 30 ~C and 6.0. DA-K strain could grow well in inorganic salt medium with aniline of 2 500 mg/L. After shaking for 96 h under the conditions of pH 6.0, 30 ℃, 180 r/min and 1 000 mg/L of aniline, the degradation rate of the aniline by DA-K could reach nearly 80%. [ Conclusion] The DA-K strain had a higher aniline degradation efficiency and actual treatment capability of the aniline wastewater, which laid foundation for establishing gene engineerincl strain.

低能离子注入诱变法选育新硫肽类抗生素166A高产菌株

目的为进一步提高产量,以小单孢菌Micromonospora sp.TMD166-MU1为出发菌株,采用低能离子注入技术,研究不同离子注入参数对菌株存活率、正负突变率的影响,通过突变株抑菌活性变势参数分析,初步探讨N+离子注入对166A生产菌所产生的诱变效果,并结合卡那霉素抗性筛选及牛津杯固体发酵高通量筛选,获得高产菌株。方法利用低能N+离子注入技术对出发菌株TMD166-MU1的孢子进行辐照诱变,以卡那霉素耐受性为选择压力,不同诱变条件处理的菌悬液涂布在含有卡那霉素培养基平板上培养后获得单菌落,并以牛津杯固体发酵高通量筛选方法进行高产菌株初筛,之后对高产菌株进行摇瓶复筛,利用高效液相法检测突变株摇瓶发酵的化学效价。结果通过研究30和60 keV能量下不同注入剂量与小单孢菌正突变率的生物学效应关系,确定了在注入能量为30 keV、注入剂量4×10^(13)ions/cm^(2)、卡那霉素抗性筛选浓度为6 mg/L条件下,可获得最高达36.33%的正突变率。复筛效价是出发菌株1.5倍以上的有4株,其中突变株IK-3的166A产量是菌株TMD166-MU1的1.8倍。结论采用低能N+离子注入诱变方式,再结合抗生素抗性筛选及牛津杯固体发酵高通量筛选的集成方法能简单、高效地获得166A高产突变菌株。

5个茯苓菌株在福建省的比较栽培试验

在福建地区对5个茯苓菌株进行段木栽培试验,比较菌核的形态、产量,并分析活性成分、营养以及重金属含量。结果表明,菌株1、4的菌核产量较高,且具有较佳形态。关键活性成分三萜含量以菌株4最高,为0.28%,多糖含量以菌株3最高,为1.37%。5个茯苓菌株富集重金属的情况不同,但检测的4种重金属含量均远低于国家限定值。综合考虑菌核产量、形态及活性成分含量等指标,菌株4为福建地区段木栽培茯苓的优良菌株。