Project description:Grifola frondosa is a valuable edible fungus with high nutritional and medicinal values. The mating systems of fungi not only offer practical strategies for breeding, but also have far-reaching effects on genetic variability. Grifola frondosa has been considered as a sexual species with a tetrapolar mating system based on little experimental data. In the present study, one group of test crosses and six groups of three-round mating experiments from two parental strains were conducted to determine the mating system in G. frondosa. A chi-squared test of the results of the test-cross mating experiments indicated that they satisfied Mendelian segregation, while a series of three-round mating experiments showed that Mendelian segregation was not satisfied, implying a segregation distortion phenomenon in G. frondosa. A genomic map of the G. frondosa strain, y59, grown from an LMCZ basidiospore, with 40.54 Mb and 12 chromosomes, was generated using genome, transcriptome and Hi-C sequencing technology. Based on the genomic annotation of G. frondosa, the mating-type loci A and B were located on chromosomes 1 and 11, respectively. The mating-type locus A coded for the β-fg protein, HD1, HD2 and MIP, in that order. The mating-type locus B consisted of six pheromone receptors (PRs) and five pheromone precursors (PPs) in a crossed order. Moreover, both HD and PR loci may have only one sublocus that determines the mating type in G. frondosa. The nonsynonymous SNP and indel mutations between the A1B1 and A2B2 mating-type strains and the reference genome of y59 only occurred on genes HD2 and PR1/2, preliminarily confirming that the mating type of the y59 strain was A1B2 and not A1B1. Based on the genetic evidence and the more reliable molecular evidence, the results reveal that the mating system of G. frondosa is tetrapolar. This study has important implications for the genetics and hybrid breeding of G. frondosa.
Project description:Proteomic analysis typically has been performed using proteins digested with trypsin because of the excellent fragmentation patterns they produce in collision induced dissociation (CID). For analyses in which high protein coverage is desirable, such as global monitoring of post-translational modifications, additional sequences can be seen using parallel digestion with a second enzyme. We have benchmarked a relatively obscure basidomycete-derived zinc metalloendopeptidase, Lys-N, that selectively cleaves the amide bond N-terminal of lysine residues. We have found that Lys-N digestion yields peptides with easily assigned CID spectra. Using a mixture of purified proteins as well as a complex yeast lysate, we have shown that Lys-N efficiently digests all proteins at the predicted sites of cleavage. Shotgun proteomics analyses of Lys-N digests of both the standard mixture and yeast lysate yielded peptide and protein identification numbers that were generally comparable to trypsin digestion, whereas the combination data from Lys-N and trypsin digestion substantially enhanced protein coverage. During CID fragmentation, the additional amino terminal basicity enhanced b-ion intensity which was reflected in long b-ion tags that were particularly pronounced during CID in a quadrupole. Finally, immonium ion peaks produced from Lys-N digested peptides originate from the carboxy terminus in contrast to tryptic peptides where immonium ions originate from the amino terminus.
Project description:Hydrophobins (HFBs) are a group of small, secreted amphipathic proteins of fungi with multiple physiological functions and potential commercial applications. In this study, HFB genes of the edible mushroom, Grifola frondosa, were systematically identified and characterized, and their transcriptional profiles during fungal development were determined. In total, 19 typical class I HFB genes were discovered and bioinformatically analyzed. Gene expression profile examination showed that Gf.hyd9954 was particularly highly upregulated during primordia formation, suggesting its major role as the predominant HFB in the lifecycle of G. frondosa. The wettability alteration profile and the surface modification ability of recombinant rGf.hyd9954 were greater than for the Grifola HFB HGFII-his. rGf.hyd9954 was also demonstrated to form the typical class I HFB characteristic-rodlet bundles. In addition, rGf.hyd9954 was shown to possess nanoparticle characteristics and emulsification activities. This research sheds light on the regulation of fungal development and its association with the expression of HFB genes.
Project description:The culinary-medicinal mushroom Grifola frondosa is widely cultivated in East Asia. In this study, the complete mitochondrial genome of G. frondosa was determined using Illumina sequencing. The circular molecule was 197,486 bp in length with a content of 25.01% GC, which was one of the largest mitochondrial genomes in the order Polyporales. A total of 39 known genes encoding 13 common mitochondrial genes, 24 tRNA genes, 1 ribosomal protein s3 gene (rps3), and 1 DNA polymerase gene (dpo) were predicted in this genome. The phylogenetic analysis showed that G. frondosa clustered together with Sparassis crispa, Laetiporus sulphureus, Wolfiporia cocos, and Taiwanofungus camphoratus. The complete mitochondrial genome reported here may provide new insight into genetic information and evolution for further studies.