Studies On The Functions Of HbSGT1 Conferring Resistance To Powdery Mildew In Hevea Brasiliensis Muell.Arg.

Powdery mildew is a kind of leaf disease with high incidence in rubber tree planting area.There is no effective way to prevent and control powdery mildew.The sequence of SGT1 gene is more conservative in eukaryotes.SGT1 is important for plant growth,development and stress signal response.The role of SGT1 in the resistance response of rubber tree powdery mildew is unclear.By using bioinformatics,molecular biology and plant pathology techniques,two rubber tree SGT1 genes,HbSGT1a and HbSGT1b,were cloned based on the sequencing of the rubber tree transcriptome;quantitative RT-PCR was used to analyze the expression patterns of HbSGT1a,HbSGT1b,HbCPK1,HbCRK1 and HbHSP90s,including various tissues,multiple hormone stimulation,drought stress treatment,mechanical wounding treatment,and powdery mildew infection;Arabidopsis thaliana transfected with HbSGT1a and HbSGT1b genes was obtained by transgenic technology;The interaction between HbSGT1a and HbHSP90.1 was verified by yeast two-hybrid system.The research results are as follows:1 HbCPKl and HbCRK1 gene were cloned,their ORF region were 1734bp and 1785 bp,encoding 577 amino acids and 595 amino acids,respectively.Bioinformatics analysis found that all contain a specific S_TKc domain.The HbCPKl and HbCRK1 proteins have no signal peptides or transmembrane domains.2 HbSGT1a and HbSGT1b genes were cloned,their ORF regions were 1086bp and 1068bp,encoding 361 and 355 amino acids,respectively.Bioinformatics analysis found that all contain a specific SGT1 domain.Subcellular localization prediction analysis found that both HbSGT1a and HbSGTlb are localized in the cytoplasm.3 Eight HbHSP90 gene family members were cloned,including HbHSP90.1,HbHSP90-3,HbHSP90-4,HbHSP90-5,HbHSP90-7,HbHSP90-8,HbHSP90-11 and HbHSP90-15.Their ORF region were 2106bp,186bp,303bp,372bp,1791bp,591bp,375bp and 435bp,respectively.The specific HATPase_c domain was found at positions 27-182 in the HbHSP90.1 sequence.Subcellular localization prediction found that HbHSP90.1 was localized in the cytoplasm.Expression analysis showed that all family members responded to the stress treatment.4 qRT-PCR analysis revealed that HbSGT1a and HbSGTIb were up-regulated after inoculation with powdery mildew.HbHSP90.1 was upregulated during hormone treatment,drought stress,mechanical wounding and powdery mildew infection.The expression of HbCPKl was the highest in flowers.The expression of HbCRK1 was the highest in leaves.HbCPKl and HbCRK1 were up-regulated in hormone treatment,drought stress,mechanical injury and powdery mildew infection.Through co-expression analysis of HbSGT1,HbHSP90,HbCPK1 and HbCRK1,it was found that all of them were up-regulated after infection by powdery mildew.5 HbSGT1a and HbSGT1b genes were located in the cytoplasm.HbSGTla and HbSGT1b were transferred to Arabidopsis plants using transgenic technology.SGT1 gene plays an important role in Arabidopsis growth and stress response.The sgtlb mutation results in phenotypes of Arabidopsis bolting earlier,loss of leaf villi,and early flowering.6 The HbSGT1 and HbHSP90.1 genes were transferred into yeast strain AH 109 by constructing a yeast two-hybrid vector.The interaction between HbSGT1a and HbHSP90.1 was confirmed by point-to-point yeast two-hybrid technique.This study lays the foundation for elucidating the resistance mechanism of HbSGT1s in powdery mildew infection.It also lays a foundation for screening resistant candidate genes and breeding rubber tree powdery mildew resistant varieties through transgenic technology.