| Auricularia cornea is the most widely distributed medicinal and edible mushroom in the world.It is the most important cultivar of the genus Auricularia in China,at present,China has become the world largest producer and exporter of A.cornea(Auricularia cornea).However,production of A.cornea in industries is on full bloom,but the basic research on the relationship between color appearance indicators and melanin is extremely weak.The most important color traits are only observed by the naked eye.It is difficult to recognize a quantitative description between color characteristics of fruiting bodies,seriously affecting the color breeding process and development of A.cornea.Therefore,on the basis of the team predicted the pigment synthesis pathway of A.cornea,this study extracted melanin from A.cornea,improved and optimized the extraction process,characterized and identified the extracted melanin,determined the structure of melanin and the physical and chemical properties in A.cornea;Quantitative and data-based evaluation of the color of A.cornea was conducted,Furthermore to analyze the link between melanin with Color characters and gel content of A.cornea;Based on the genome of the A.cornea and the key enzyme gene G11815 of the annotated pigment synthesis obtained by the team,the stable expression of the reference gene was screened for reference.Based on the genome of the A.cornea and the key enzyme gene G11815 of the annotated pigment synthesis obtained by the team,using qRT-PCR,the stable expression of the reference gene was screened for reference.The expression of G11815 in brown and white A.cornea was determined,to verify the mechanism of color synthesis of white A.cornea.Verified the synthesis mechanism of white A.cornea.The aim is to provide a new way for exploring the relationship between color and internal pigment and directional color breeding of A.cornea,and to provide scientific support for cultivating more new varieties of edible fungi with different colors.The main test results are as follows:1.Extraction,characterization,physical and chemical properties of melanin from A.cornea(1)The optimal extraction conditions of melanin from A.cornea were as follows:concentration of sodium hydroxide solution 1.5 mol/L,ratio of material to liquid 1:30(g/mL),extraction time 1 h.(2)The extracted melanin was analyzed,the results showed that melanin was alkalophilic and hydrophobic,and it was insoluble in acid solution and organic solvent;melanin had the maximum absorption peak at 210 nm;melanin structure was catechol.(3)Stability:The consumption rate of melon melanin after passing through 100°C water bath for 1 h was 25.20%,and the consumption rate under incandescent light treatment was as high as 39.81%.The thermal stability of A.cornea was higher than that of light stability.Antioxidant:When the concentration of melanin in A.cornea is above 0.5mg/mL,the DPPH free radical scavenging rate can reach more than 60%,and the IC50 is 0.46mg/mL.2.The correlation between melanin content and fruit body color and gel content of A.cornea(1)The black standard(L~*=0,a~*=0,b~*=0)was used as the base standard,The results showed that the color of the ventral surface of the A.cornea was black reddish and blue,and the brightness was bright;the back color is black,green,blue,and bright.The Moquent mathematical model was used to analyze the color of the fruiting body of A.cornea.It was found that the color traits of A.cornea had a certain continuity.(2)The melanin content of A.cornea was significantly negatively correlated with the L~*value of the ventral surface color of the fruit body,and the correlation coefficient was-0.506.There was a significant negative correlation with the gel content,and the correlation coefficient was-0.714.(3)The highest-yield melanin strain AP17 was screened.3.Expression analysis of internal reference gene screening and key enzyme gene G11815 of pigment synthesis(1)Three strains of brown A.cornea and three white A.cornea were selected,and?-tubulin(?-TUB),glucose phosphate mutase(PGM),phosphoglucose isomerase,(PGI)plasma membrane proton ATPase(H~+-ATPase)were selected from the transcriptome data of A.cornea.and designed primers.The optimal internal reference gene was PGM in brown A.cornea,and the most suitable internal reference gene in white A.cornea was H~+-ATPase.(2)The expression of glutaminamide transferase gene G11815,a key enzyme in the synthesis of A.cornea pigment,was determined by using the selected reference gene as a reference.The results showed that the relative expression level of G11815 in brown A.cornea was 380 times that of white A.cornea.It is verified that G11815(glutamine aminotransferase)is the key enzyme gene of pigment synthesis in A.cornea,the reason why white A.cornea fruiting body appears white is that the expression of G11815 is inhibited or the expression is absent. |
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