ABSTRACT
The enoki mushroom (Flammulina filiformis) is one of the most important and popular edible mushrooms commercially in China. However, traditional mushroom cultivar identification is challenging due to poor accuracy, heavy workloads, and low reproducibility. To overcome this challenge, we developed a method for identifying F. filiformis strains using multiple nucleotide polymorphism sequencing (MNP-seq). This involved screening 179 universal MNP markers based on whole-genome sequencing data, constructing an MNP sequence library, and performing multiplex PCR amplification and high-sequencing. We further screened 69 core MNP markers and used them to build a neighbor-joining (NJ) phylogenetic tree of 232 cultivated and wild strains. Our analysis showed that all cultivars could be accurately separated by computing genetic similarity values and that the cultivars could be separated into 22 distinct evolutionary pedigrees. The specific value of genetic similarity can be used as the standard to distinguish F. filiformis cultivars, however, it needs to be comprehensively defined by the additional phenotype and biological characteristics of those strains in the future work.
ABSTRACT
Auricularia cornea Ehrenb., previously named A. polytricha (Mont.) Sacc, has become one of the most widely cultivated mushrooms in China. Considerable research has been conducted on its cultivation, pathogen identification, proteomics, and more. However, to the best of our knowledge, no studies have been performed on reference-gene validation in this species. Formerly, reference genes were selected for their expression levels only relied upon from others species, owing to the fact that the gene stability in this species is unknown. In this study, nine candidate genes, including tubulin alpha-1A chain (TUBA1A), ß-tubulin (Btu), phosphoglucomutase (Pgm), actin 1 (Act1), protein phosphatase 2A regulatory subunit (PP2A), polyubiquitin (UBQ), glyceraldehyde-3-phosphate dehydrogenase (Gapdh), 18S ribosomal protein (18S) and 28S ribosomal protein (28S), were evaluated among different strains and developmental stages. Four algorithms (i.e., geNorm, NormFinder, BestKeeper and RefFinder) were used to analyze candidate genes. The results revealed that UBQ was the most stable reference gene, while 18S was the least stable. Despite these results, the candidate genes were largely inadequate and only two were considered suitable. Based on candidate gene stability, PP2A and UBQ were identified as a set of usable interior control genes for future analyses in this species. This is the first systematic study conducted for selecting reference genes in A. cornea, and lays the foundation for identifying genes and quantifying gene expression in this species.
Subject(s)
Agaricales/genetics , Gene Expression Profiling , Gene Expression , Genes, Fungal , Real-Time Polymerase Chain Reaction/methodsABSTRACT
OBJECTIVE: To investigate the genetic diversity of Ganoderma cultivars provided for the genuineness study, germ-plasm resource identification, genetic relationship study, breeding, introduction and cultivante of Ganoderma strains. METHOD: With the software of NTSYSpc 2. 1, 24 materials, of G. lucidum and G. sinense, were studied using AFLP to construct the dendrogram. RESULT: There were 177 polymorphic bands with 14 primer combinations. And all materials could be identified with AFLP. CONCLUSION: There actually existed much genetic diversity at the molecular level among the germplasm resources of Ganoderma strains, and all the strains were clustered into G. lucidum group and G. sinense group at the similarity coefficient 0. 676.
