In order to enhance the utilization efficiency, reduce the environmental pollution of traditional chemical treatment and the agriculture waste incineration; we studied the ligninase production by Coprinus comatus, Aspergillus niger and Trichoderma reesei through the plate screening. The results showed that C. comatus mixed culture with T. reesei have a good compatibility and higher yields of Laccase. On the basis of this pre-experiment, we studied the optimal conditions of mixed culture for enzyme production. Under the optimal conditions: the inoculation proportion of C. comatus and T. reesei (5:2), the interval of time (12 h), the temperature 260C, the shake rotation speed 150 r/min, fermented for 3 days, the Laccase activity reached 3267.1 U/mL, increased by 106% contrasted with single culture of C. commatus.
Coprinus ; metabolism ; Culture Techniques ; methods ; Fermentation ; Oxygenases ; biosynthesis ; Plant Stems ; metabolism ; Trichoderma ; metabolism ; Zea mays ; metabolism

Trichoderma spp. is a kind of filamentous fungi with important biocontrol value. Twelve strains of Trichoderma spp. were isolated from the soils of different types of crops in Shaoxing, Zhejiang and Foshan, Guangdong. The antagonistic resistance to Fusarium oxysporum was compared by plate confrontation test. The further analysis of volatile secondary metabolites for two strains were carried out using HS-SPME-GC-MS analysis. The results showed that T. asperellum ZJSX5003 and GDFS1009 had fast growth ability, and the inhibition effects on F. oxysporum were 73% and 74% respectively. Six identical volatile metabolites were detected as follows 2-Methyl-1-propanol, 3-Methyl-1-butanol, 3-Methyl-3-buten-1-ol, Acetyl methyl carbinol, Butane-2,3-diol and 6-n-pentyl-2H-pyran-2-one (6-PAP). Among them, 6-PAP was validated to have a higher inhibitory effect on F. oxysporum in vitro. This study will provide basis for the development of biocontrol agents with metabolites of Trichoderma, such as 6-PAP.
Antibiosis ; Antifungal Agents ; pharmacology ; Fusarium ; drug effects ; physiology ; Gas Chromatography-Mass Spectrometry ; Trichoderma ; chemistry ; metabolism

Ethylene is the most widely used petrochemical feedstock globally. The development of bio-ethylene is essential due to limited fossil fuels and rising oil prices. Bio-ethylene is produced primarily by the dehydration of ethanol, but can alternatively be directly produced from ethylene biosynthesis pathways in plants, algae, or microorganisms by using cheap and renewable substrates. This review addressed the biosynthesis of ethylene in plants and microorganisms, the characterization of key enzymes, genetic engineering strategies for ethylene biosynthesis in microorganisms, and evaluated its perspective and successful cases toward the industrial application. The direct production of bio-ethylene from a biological process in situ is promising to supplement and even replace the petrochemical ethylene production.
Ethylenes ; biosynthesis ; Industrial Microbiology ; methods ; Metabolic Engineering ; methods ; Plants ; genetics ; metabolism ; Saccharomyces cerevisiae ; metabolism ; Synechocystis ; genetics ; metabolism ; Trichoderma ; metabolism

After the cell enters into its programmed cell death, xylanases from grass plants gradually matured through its N-terminal and C-terminal sequence been cut by acid proteases several times. They could not be expressed by conventional protein expression system. Search the GenBank database, xynIII from a mutant of T. reesei QM9414(ATCC26921)was found. It is similar to grass plants' xylanase in their families and structures. It couldn't express in T. reesei QM9414, but its gene exist in genomic DNA as one copy. Through overlap-PCR method, 4 exons of xynIII were cloned, sequenced, spliced, and the whole cDNA of mature xynIII was acquired. The cDNA was inserted into pETBlue-2 vector and transformed into E. coli DE3 pLacI cell. Xyn III could be expressed in the transformed cell under the conditions of 37 degrees C, 1 mmol/L IPTG induced for 3h. Low temperature (15 degrees C), long time(64h) induction(0.2 mmol/L IPTG) could enhance xynIII activity.
Cloning, Molecular ; DNA, Complementary ; chemistry ; Endo-1,4-beta Xylanases ; genetics ; metabolism ; Polymerase Chain Reaction ; methods ; Trichoderma ; genetics

From the fungus Trichoderma sp., we isolated seven novel 18-residue peptaibols, neoatroviridins E-K (1-7), and six new 14-residue peptaibols, harzianins NPDG J-O (8-13). Additionally, four previously characterized 18-residue peptaibols neoatroviridins A-D (14-17) were also identified. The structural configurations of the newly identified peptaibols (1-13) were determined by comprehensive nuclear magnetic resonance (NMR) and high-resolution electrospray ionization tandem mass spectrometry (HR-ESI-MS/MS) data. Their absolute configurations were further determined using Marfey's method. Notably, compounds 12 and 13 represent the first 14-residue peptaibols containing an acidic amino acid residue. In antimicrobial assessments, all 18-residue peptaibols (1-7, 14-17) exhibited moderate inhibitory activities against Staphylococcus aureus 209P, with minimum inhibitory concentration (MIC) values ranging from 8-32 μg·mL^-1. Moreover, compound 9 exhibited moderate inhibitory effect on Candida albicans FIM709, with a MIC value of 16 μg·mL^-1.
Peptaibols/chemistry* ; Trichoderma/metabolism* ; Tandem Mass Spectrometry/methods* ; Anti-Infective Agents/pharmacology* ; Spectrometry, Mass, Electrospray Ionization/methods*

It was expected that recombinant Aspergillus niger glucose oxidase could be expressed in Trichoderma reesei with stable activity. T. reesei CBHI promoter--CBHI ss. gene--A. niger glucose oxidase gene--T. reesei CBHI terminator--A. nidulans gpd promoter--E. coli Hygromycin B phosphotransferase gene--A. nidulans trpC terminator--pUC19 (pCBHGOD) vector was constructed in E. coli DH5alpha by PCR application and gene cloning methods. T. reesei QM9414 protoplast was transformed by T. reesei CBHI promoter-CBHI ss. Gene--A. niger glucose oxidase gene--T. reesei CBHI terminator-A. nidulans gpd promoter--E. coli Hygromycin B phosphotransferase gene--A. nidulans trpC terminator linear DNA fragment (CBHGOD fragment) that was made by digestion of pCBHGOD with Kpn I. T. reesei mutant clone with homologous recombinant A. niger glucose oxidase gene was selected by PCR method. Recombinant glucose oxidase was produced by mutant T. reesei strain under induction of wheat straw for 5 days. Recombinant glucose oxidase molecular mass was showed the same as native A. niger glucose oxidase standard from Sigma company by Western blot analysis. Recombinant glucose oxidase activity was 25u/mL in medium. The yield was 0.5 g/L in comparison with Sigma company glucose oxidase standard. There was no recombinant GOD degradation during Trichoderma reesei cultivation that was showed in Western blot analysis. Trichoderma reesei has capability to be a new recombinant host for Aspergillus niger GOD production.
Aspergillus niger ; enzymology ; genetics ; Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Fungal Proteins ; genetics ; metabolism ; Glucose Oxidase ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Trichoderma ; genetics ; metabolism

Antagonistic mechanisms of Trichoderma viride M3, Tv04-2, and T. harzianum ThB, were studied against Phytophthora nicotianae, the pathogen of stem blight disease on Schizonepeta tenuifolia by dual-culture, hydrolase activity, volatile and nonvolatile substances. Results indicated that competitive, mycoparasitism and antagonism were the antagonistic mechanisms of three Trichoderma spp. against P. nicotianae. Hydrolase activity showed that M3 was the highest for beta-1, 3-glucanases activity while ThB was the highest for proteases activity among the three T. strains, and they could produce volatile and non-volatile substances, also.
Fungal Proteins ; metabolism ; Hydrolases ; metabolism ; Lamiaceae ; parasitology ; Peptide Hydrolases ; metabolism ; Pest Control, Biological ; Phytophthora ; microbiology ; physiology ; Plant Diseases ; parasitology ; Trichoderma ; enzymology ; physiology

OBJECTIVETo identify the endophytic fungal strain PR35 separated from Paeonia delavayi and study chemical constituents of its secondary metabolites.

METHODThe fungal strain PR35 was identified by morphological observation and ITS rDNA sequence analysis. Various chromatographic methods were adopted to separate and purify its secondary metabolites, and their structures were identified by physiochemical properties and spectral data

RESULTThe fungal strain PR35 was identified as Trichoderma longibrachiatum. Five compounds were separated from fermentation products of fungal strain PR35 and identified as 1-(2,6-dihydroxyphenyl)-3-hydroxybutan-1-one (1), 1-(2,6-dihydroxypheny) propan-1-one (2), 1-(2,4,6-trihydroxyphenyl) butan-1-one (3), 4-methoxy-1-naphthol (4), and cerevisterol (5). Among them, compounds 1-3 showed notable antifungal activities against Botrytis cinerea, Fusarium avenaceum and Hormodendrum compactum.

CONCLUSIONThe endophytic fungus T. longibrachiatum was separated from the plant P. delavayi for the first time. Five compounds were first separated from endophytic fungus of P. delavayi. Among them, compound 4 was separated from microbial fermentation products for the first time.

DNA, Fungal ; genetics ; DNA, Intergenic ; genetics ; Endophytes ; classification ; genetics ; isolation & purification ; metabolism ; Paeonia ; microbiology ; Phylogeny ; Trichoderma ; classification ; genetics ; isolation & purification ; metabolism

Complete mannanase gene with two introns was cloned from Trichoderrna reesei by PCR. The two introns were then removed by overlap extension PCR. The gene encoding the mature mannanase protein was inserted into the expression vector pPIC9K, downstream of a alpha-factor signal peptide sequence. The resultant recombinant vector was named pM242. After linearized with Sac I , pM242 was transformed to Pichia pastoris GS115 by electroporation. After screening, the recombinant strain Gpmf25 that expresses the secretory protein at high level was obtained. The activity of the recombinant mannanase reached 12.5 IU/mL. Optimum pH and temperature for the recombinant enzyme were 5.0 and 80 degrees C, respectively. The enzyme was stable at pH 5.0-6.0 and maintained over 50% of original activity after incubation at 70 degrees C for 30 min.
Fungal Proteins ; biosynthesis ; genetics ; Hydrogen-Ion Concentration ; Pichia ; genetics ; metabolism ; Recombinant Proteins ; biosynthesis ; genetics ; Temperature ; Trichoderma ; enzymology ; genetics ; beta-Mannosidase ; biosynthesis ; genetics

OBJECTIVETo obtain transgenic Pinellia ternata plants resistant to fungus by transfer Chitinase and beta-1,3-Glucanase gene from Trichoderma harzianum.

METHODUsing hygromycin phosphotransferase as the selection marker, the Chitinase gene (ech42), beta-1,3-Glucanase gene (gluc78) and both gene pCAMBIA(ech42 + gluc78) driven by CaMV35S promoter were transferred into P. ternata callus via Agrobacterium-mediated transformation.

RESULTPCR results confirmed that the regenerants were identified to be transgenic lines and the RT-PCR results confirmed that foreign genes construction were transfer to mRNA. Two foreign genes were inherited stably to T5 generation according to PCR results of the lines.

CONCLUSIONThe results showed that chitinase gene ech42 and beta-1, 3-glucanase gene gluc78 respectively or together introducing and co-integrating into P. ternata

Agrobacterium tumefaciens ; genetics ; metabolism ; Chitinases ; genetics ; metabolism ; Fungal Proteins ; genetics ; metabolism ; Gene Expression Regulation, Plant ; Gene Transfer Techniques ; Genetic Vectors ; genetics ; metabolism ; Glucan 1,3-beta-Glucosidase ; genetics ; metabolism ; Pinellia ; genetics ; metabolism ; Transformation, Genetic ; Trichoderma ; enzymology