Preliminary Study On The Function Of Z91 Gene In The Interaction Between Sclerotinia Sclerotiorum And Fungal DNA Virus SsHADV-1

Sclerotinia sclerotiorum is a worldwide distributed of plant pathogenic fungus,which infects a variety of crops such as rapeseed and soybean,causing severe economic losses.Mycovirus SsHADV-1 virus can confer hypovirulence to S.sclerotiorum,after infected by the virus,S1 showed a serious recession,including slow growth rate,delayedsclerotial formation,restrained and uneven pigment production and hypovirulence.Hence,SsHADV-1 has the potential for the prevention and control of diseases.S.sclerotiorum also is counterproductive with SsHADV-1,as well as gets rid of the virus in the process of cultivation,which makes using of mycovirus for crop disease control a potential risk.Therefore,further study of molecular mechanism in the interaction of S.sclerotiorum and SsHADV-1 is needed.In order to clarify the interaction mechanism between SsHADV-1 and S.sclerotiorum,gene expression profiles of S.sclerotiorum before and after virus infection were analyzed using Solexa(RNA-seq)technique.Based on the digital gene expression profile(DGE),we obtained a gene named Z91 which declined by 16 times after virus infection.The function of this gene in the interaction of S.sclerotiorum and SsHADV-1 was studied using gene silence and over expression technology.The main results are listed as following:1.Z91 gene is composed of 1498 bp containing three exons and two introns,encoding a protein of 410 amino acids.The predicted protein has two conserved domains of Pall and filament_head.qRT-PCR experiment showed that the expression level of Z91 gene declined by 10 times after virus infected 1-3 h,The results showed the gene may be involved in virus infection of S.sclerotiorum.2.Over expression and silencing vector of Z91 were constructed,and then transformed to S.sclerotiorum strain Ep-1PNA367 by ATMT method.Compared to strain Ep-1PNA367,the expression level of Z91 declined in the silenced transformants,and increased in the over expressed transformants.3.Biological experiments suggested that over expression of Z91 gene in S.sclerotiorum has no significant impact on mycelial growth and virulence,while the silenced transformants have varying degrees of decreases in mycelial growth rate and pathogenicity,producing no sclerotia and sensitivity to high salt and high osmotic pressure.It suggested that Z91 gene is involved in mycelial growth,sclerotial formation and pathogenicity and plays important roles in resistance to stress.4.SsHADV-1 particles could infect over expressed transformants and silenced transformant of Z91 gene,SsHADV-1 replication was not affected significantly in the over expressed transformants,but in silenced transformants,SsHADV-1 replication was significantly suppressed,and the genome DNA of SsHADV-1 could only be detected by PCR anplification.It may indicate that the Z91 gene plays an important role in interaction of S.sclerotiorum and SsHADV-1.These results primarily indicated the function of Z91 gene in growth and development,pathogenicity and Z91 gene plays an important role in the accumulation of SsHADV-1 in S.sclerotiorum,but further research is necessary to the interaction mechanism.