In this study, we employed a combination of genome mining and heteronuclear single quantum coherence (HSQC)-based small molecule accurate recognition technology (SMART) technology to search for fernane-type triterpenoids. Initially, potential endophytic fungi were identified through genome mining. Subsequently, fine fractions containing various fernane-type triterpenoids were selected using HSQC data collection and SMART prediction. These triterpenoids were then obtained through targeted isolation and identification. Finally, their antifungal activity was evaluated. As a result, three fernane-type triterpenoids, including two novel compounds, along with two new sesquiterpenes and four known compounds were isolated from one potential strain, Diaporthe discoidispora. Their structures were elucidated through analysis of high-resolution electrospray ionization mass spectrometry (HR-ESI-MS) and nuclear magnetic resonance (NMR) spectroscopic data. The absolute configurations were determined using single-crystal X-ray diffraction analysis and electron capture detector (ECD) analysis. Compound 3 exhibited moderate antifungal activity against Candida albicans CMCC 98001 and Aspergillus niger.
Triterpenes/isolation & purification* ; Antifungal Agents/isolation & purification* ; Molecular Structure ; Candida albicans/drug effects* ; Ascomycota/genetics* ; Magnetic Resonance Spectroscopy ; Aspergillus niger/drug effects* ; Genome, Fungal ; Microbial Sensitivity Tests

Filamentous fungi represent an important group of eukaryotic microorganisms with diverse ecological functions and ubiquitous distribution in various ecosystems. Among them, many species are closely associated with agriculture, functioning as major plant pathogens that cause yield losses and produce mycotoxins to compromise both the quality and safety of agricultural products. In recent years, the CRISPR/Cas system has emerged as a powerful and programmable genome editing tool, and it has been extensively applied to the genetic study of plant pathogenic fungi. This technology has greatly facilitated the investigation of pathogenic mechanisms, mycotoxin biosynthetic pathways and key gene functions, antifungal resistance, and rapid pathogen detection. This review summarizes the development of CRISPR/Cas systems and the key strategies for their application in plant pathogenic fungi and makes an outlook on the practical deployment. With the continuous advancement of gene editing technologies, emerging fungal-adapted editing systems hold great promise for advancing functional genomics and enabling innovations in disease-resistant breeding and sustainable crop protection.
Gene Editing/methods* ; Fungi/pathogenicity* ; CRISPR-Cas Systems/genetics* ; Plant Diseases/microbiology* ; Plants/microbiology* ; Genome, Fungal/genetics*

Aspergillus niger, as an important industrial fermentation strain, is widely applied in the production of organic acids and industrial enzymes. With the development of diverse omics technologies, the data of genome, transcriptome, proteome and metabolome of A. niger are increasing continuously, which declared the coming era of big data for the research in fermentation process of A. niger. The data analysis from single omics and the comparison of multi-omics, to the integrations of multi-omics based on the genome-scale metabolic network model largely extends the intensive and systematic understanding of the efficient production mechanism of A. niger. It also provides possibilities for the reasonable global optimization of strain performance by genetic modification and process regulation. We reviewed and summarized progress in omics research of A. niger, and proposed the development direction of omics research on this cell factory.
Aspergillus niger ; genetics ; Fermentation ; Genome, Fungal ; Metabolic Networks and Pathways ; Metabolome ; Proteome ; Transcriptome

Chromosomal integration of heterologous genes or pathways is preferred over the use of episomal plasmids for its inherently stability and thus more desirable in the industrial setting. However, the position of integration of heterologous genes in the genome influences the expression levels. In combination of high throughput transformation of the Yeast Knock-out Collection (YKO) and FACS analysis, the position effect on heterologous reporter gene gfp was identified across the whole genome in yeast. In total 428 high-expressed sites and 444 low-expressed sites were spotted, providing massive data to analyze patterns and reasons for region dependency of gene expression on the genome-wide scale.
Gene Expression Regulation, Fungal ; Gene Knock-In Techniques ; Genes, Reporter ; Genome, Fungal ; Saccharomyces cerevisiae ; genetics

Codon usage bias is an important characteristic of genetic information transfer in organisms. Analysis of codon usage bias of different species is important for understanding the rules on genetic information transfer. The previous method for analysis of codon usage bias is mainly based on genomic data. However, this method is greatly limited, because the genome sequences of higher organisms are still not available up to now. In this study, we found that we could obtain the same optimal codons of Ganoderma lucidum (Curtis: Fr.) P. Karst based on its whole genomic data or large-scale transcriptomic data from its liquid-cultured hyphae, primordium and fruiting body, separately. This result indicated the feasibility to understand the codon usage bias based on the large-scale transcriptomic data. By calculating the proportion of rare codons of Escherichia coli and Saccharomyces cerevisiae in 26 terpene synthases (TS) of G. lucidum, we found that the rare codons of S. cerevisiae have a higher proportion in TS genes, while the rare codons of E. coli have relatively lower, suggesting that the TS genes of G. lucidum are possibly more difficult to be expressed in S. cerevisiae than in E. coli. Chemical synthesis of TS genes according to the yeast optimal codons will be an effective way to solve the problem on the mismatch of gene codon bias between the foreign genes and the host strain.
Codon ; Escherichia coli ; Genome, Fungal ; Reishi ; genetics ; Saccharomyces cerevisiae ; Transcriptome

OBJECTIVETo establish the EST-SSR marker system for Cordyceps by using ESTs of C. bassiana and C. militaris.

METHODThe ESTs of Cordyceps were downloaded from the public database of NCBI, and the redundant ESTs with low quality were removed. The EST-SSR primers were designed by Sequece Seiner 1. 2. And the primers were screened through PAGE-Electrophoresis.

RESULTThe 4 556 non-redundant ESTs which from C. bassiana with total length of 2 953 173 bp were selected. 718 EST-SSRs distributed in 616 ESTs were totally screened out, accounting for 15.8% of the non-redundant ESTs. It was discovered that the average distance of EST-SSSR was 1/4 096 bp in EST-SSRs distribution of C. bassiana. Trinucleotide repeats were the most abundant types with 419 repeated sequences. Regarding to C. militaris, totally 1 363 non-redundant ESTs were acquired, from which 1 117 EST-SSRs were screened, and rate of SSR sites in ESTs was 81.95%. The leading motif of SSR was nucleotide A. The 50 pairs of EST-SSR primers were designed according to the ESTs of C. bassiana, and preliminary test showed the 34 pairs of primers amplified clear fragments,accounting for 68% of all primers. Furthermore, the 39 of the 40 pairs of primers from the ESTs of C. militaris were found to be amplified as the clear fragments, accounting for 97.5%. The phylogenetic analysis revealed that different anamorph of Cordyceps spieces were divided into four branches.

CONCLUSIONThe EST-SSR of Cordyceps had comparably higher utility value. The EST-SSR markers developed from ESTs of C. bassiana and C. militaris had well transferability in Cordyceps. And it was suggested that the EST-SSR markers should be an easy and effective way to assay molecular genetic structure of Cordyceps.

China ; Cordyceps ; classification ; genetics ; DNA Primers ; DNA, Fungal ; genetics ; Databases, Nucleic Acid ; Expressed Sequence Tags ; Genetic Markers ; genetics ; Genome, Fungal ; genetics ; Microsatellite Repeats ; genetics ; Phylogeny ; Polymorphism, Genetic ; Repetitive Sequences, Nucleic Acid ; genetics

Aspergillus niger is an important industrial workhorse with extensive application in the sectors of industrial enzymes, heterogeneous proteins, organic acids and etc. The disclosure of its genomic sequence to the public brought the study of A. niger into the post-genomic era. Diverse omic data are being produced massively and rapidly, which largely upgrades our understanding to the hyperproduction mechanism of A. niger to a systems and molecular level. At meanwhile, its genetic operating system is becoming mature, which enables genome-scale genetic perturbation within A. niger. In conclusion, we are on the right way to redesign and engineer A. niger to an omnipotent cellular factory.
Aspergillus niger ; genetics ; metabolism ; Biotechnology ; methods ; Enzymes ; genetics ; secretion ; Gene Expression Regulation, Fungal ; Genes, Fungal ; Genome, Fungal ; Protein Biosynthesis ; genetics ; Recombinant Proteins ; secretion ; Transcription, Genetic

Inefficient degradation of lignocellulose is one of the main barriers for the utilization of renewable plant biomass for biofuel production. The bottleneck of the biorefinery process is the generation of fermentable sugars from complicated biomass polymers. In nature, the main microbes of lignocelluloses deconstruction are fungi. Therefore, elucidating the mechanism of lignocelluloses degradation by fungi is of critical importance for the commercialization of lignocellulosic biofuels. This review focuses on the progress in lignocelluloses degradation pathways in fungi, especially on the advances made by functional genomics studies.
Biofuels ; Fungi ; genetics ; metabolism ; Genetic Engineering ; Genome, Fungal ; genetics ; Industrial Microbiology ; Lignin ; metabolism

Genes, Synthetic ; Genetic Engineering ; Genome, Fungal ; Mycoplasma mycoides ; genetics ; Synthetic Biology

Plasmid-carrying Saccharomyces cerevisia (W303-1B[pYeDP60/G/ADS]) and genome-transformed S. cerevisia (W303-1B[rDNA:ADS]), both harboring amorpha-4,11-diene synthase (ADS) gene were constructed to investigate the production of amorpha-4,11-diene. The recombinant plasmid pYeDP60/G/ADS that harbors the ADS gene was transformed into S. cerevisiae W303-1B, resulting in the engineered yeast W303-1B[pYeDP60/G/ADS], which contains multi-copies of the plasmid. The ADS gene expression cassette was obtained by PCR amplification of the pYeDP60/G/ADS template, and then introduced into S. cerevisiae W303-1B to obtain the engineered yeast W303-1B[rDNA:ADS], in which the ADS gene was integrated into the rDNA locus of the yeast genome through the homologous recombination. GC-MS analysis confirmed that both of the engineered yeasts could produce amorpha-4,11-diene. Moreover, the amorpha-4,11-diene yield of W303-1B[pYeDP60/G/ADS] was higher than that of W303-1B[rDNA:ADS]. Southern blot analysis showed that there is only one copy of ADS gene in the genome of W303-1B[rDNA:ADS]. It implied that the amorpha-4,11-diene yield can be improved by increasing the ADS gene copies.
Alkyl and Aryl Transferases ; genetics ; metabolism ; DNA, Ribosomal ; genetics ; Fermentation ; Gas Chromatography-Mass Spectrometry ; methods ; Genetic Engineering ; methods ; Genome, Fungal ; genetics ; Plasmids ; Saccharomyces cerevisiae ; genetics ; metabolism ; Sesquiterpenes ; metabolism ; Transformation, Genetic