Study On Genetic Property Of The Color Mutant Of Flammulina Velutipes

Flammulina velutipes, also known as snake butter, is a fungus belonging to Flammulina, Physalacriaceae, Agaricales, Agaricomycetes,Basidiomycota. It is a famous edible fungus. The fruiting body of wild Flammulina velutipes in its natural state is yellowish-brown. Currently,there is pure white and brown Flammulina velutipes commercially cultivated, where pure white Flammulina velutipes is mutant. Although the color of Flammulina velutipes is one of the important commercial qualities, few researches are conducted on the ingredients of pigment and its synthesis pathway as well as the mechanism of how the white mutant strain is generated. Based upon the extraction and identification of pigment of brown Flammulina velutipes, the differences of key enzyme activities in terms of pigment biosynthesis of brown and white Flammulina velutipes are compared, and transcriptome sequencing is conducted on the brown and white Flammulina velutipes with the second-generation high-throughput sequencing as platform. The main research results are as follows:(1)With the use of alkali-solution and acid-isolation property of pigment,the pigment of brown Flammulina velutipes is extracted. UV-vis and infrared spectral characterization indicated that it has similar structure of synthetic tyrosine melanin, belonging to the group of 3,4-dihydroxy phenylalanine(DOPA) melanins;(2)Superoxide dismutase(SOD) activity of brown Flammulina velutipes is significantly lower than that of white Flammulina velutipes, yet its tyrosinase activity is significantly higher than that of white mutant;(3)A total of 967348960 nt data are output through high-throughput sequencing. When results are assembled, a total of Unigene 29490 are obtained, with overall length as 41,412,960 nt and average length as1404 nt. With functional annotation of Unigene, there are 21247, 6985,13260, 13234, 10062 and 7310 Unigene respectively in database NR,Swiss-Prot, KEGG, COG and GO, with a total of 31397 Unigene in all the annotations. CDS is predicted. There are 21,239 CDS after alignment to protein pool. There are 2,244 CDS that can be predicted, and in a total of 23,483, there are 1,442 SSR;(4)With the analyses of Unigene differential expression and KEGG metabolic pathway, it is concluded that tyrosinase gene plays a key role in the synthesis pathway of melanin and that the tyrosinase genes playing a key role in up-regulation are encoded as CL520 contig1, CL1942 contig2,CL1942 contig3 and CL1942 contig4;(5)Through Real time RT-PCR combined with semi-quantitative RT-PCR technology, differential expressions of tyrosinase genes between brown and white Flammulina velutipes are verified. The results are consistent with those from transcriptome sequencing analysis.