肺形侧耳退化与基因组DNA 甲基化关系的探讨

Relationship Between Degradation of Pleurotus pulmonarius and Genomic DNA Methylation

以肺形侧耳(Pleurotus pulmonarius)菌株秀2、秀t 2、秀tz 2为供试菌株,通过拮抗实验、ISSR和RAPD多态性分析研究3个供试菌株间的亲缘关系;比较3个菌株在菌丝生长速度、产量以及基因组DNA甲基化率三个方面的差异,以揭示菌株退化与基因组DNA甲基化的关系;通过5-氮杂胞苷(5-azacytidine,5-azaC)去甲基化处理菌株后,检测其基因组甲基化率,并将去甲基化处理菌株继代4次后测定甲基化率和菌丝生长速度。结果表明:秀t 2与秀tz 2、秀2与秀tz 2、秀t 2与秀2均无拮抗反应,ISSR和RAPD扩增产物经过分析后得出3个菌株亲缘关系接近;与秀t2相比,秀2、秀tz2的菌丝生长速度和第一潮产量均显著提高,基因组DNA甲基化率均显著降低。经过20 mmol·L^(-1 )5-azaC去甲基化后,与对照组(CKF1)相比,处理组(20 F1)基因组DNA甲基化率显著降低、菌丝生长速度显著提高,推测肺形侧耳菌株退化与其基因组DNA甲基化存在联系。5-azaC处理肺形侧耳,其基因组甲基化率会降低,经过4次继代后,DNA甲基化率会逐步提高,表明继代培养可能引起菌株退化。

In order to study the relationship between strain degradation and genomic DNA methylation in Pleurotus pulmonarius,three related strains of P.pulmonarius,Xiu 2,Xiu t 2 and Xiu tz 2,were first verified for their affinity through antagonistic experiment,ISSR and RAPD polymorphism analysis,and then compared for mycelial growth rate,yield and genomic DNA methylation rate.In addition,another P.pulmonarius strain Xiu 57 was demethylated by 5-azacitidine(5-azaC),subcultured successively for four passages,and then measured for mycelial growth rate and genomic DNA methylation rate.The results showed that there was no antagonistic reaction between Xiu 2,Xiu t 2 and Xiu tz 2,and they had a close genetic relationship as indicated by ISSR and RAPD markers.Compared with Xiu t 2,Xiu 2 and Xiu tz 2 showed a significantly higher mycelial growth rate and first flush yield,but a significantly lower genomic DNA methylation rate.Upon demethylation with 20 mmol·L^(-1) 5-azaC,Xiu 57 showed a significant decrease in genomic DNA methylation rate in parallel with a significant increase in mycelial growth rate,suggesting a relationship between strain degradation and methylation of genomic DNA.After four passages of subculture,the genomic DNA methylation rates of both the 5-azaC treated group and the blank control increased,suggesting that subculture may cause strain degeneration.