A Preliminary Study On The AtSb3Gene With MATH Domains From Arabidopsis

MATH domain (Meprin and TRAF Homology Domain) protein consisting of7anti-parallel β-helix can mediated protein-protein interaction. Six genes containing MATH domain were cloned from the Brassica napus genome,one of these genes was named BnD12. To reveal the function of BnD12gene, a gene named AtSb3was screened from Arabidopsis thaliana genome,which also contains MATH domain shared an high homology with BnD12gene, and the function of AtSb3gene was unknown.Such as bioinformatics, molecular biology and reverse genetics were used to study the AtSb3gene function.The results are as follows:1. Bioinformatical analysis showed that, AtSb3gene contains7exons and6introns respectively, with a1056bp full length CDS.The estimated molecular weight is39,826D and the isoelectric point were9.21, it encodes351aa which contains two MATH domains. The structure of N-terminal has a signal peptide and a predicted transmembrane domain, and supposed to be a hydrophilic protein and secreted protein.2. Semi-quantitative RT-PCR was used to analyze the expressed site of AtSb3gene in root, stem, leaf, flower, silique, it turned out that AtSb3gene was expressed specifically in root.3.1690bp upstream of the AtSb3gene as promoter region had been cloned, According to the promoter base sequence analysis, the promoter contains some root expression elements and resistance related expression elements.The promoter and GUS fusion expression vector pAtSb3pro101::GUS was constructed. Floral-dip method was used to obtain Arabidopsis thaliana transgenic lines. GUS histochemical staining demonstrated that the gene was expressed specifically in radicle and mature root,while highly expressed in root cap and elongation zone of root tip in mature root, whereas no expression between meristem and elongation zone.4. AtSb3gene two homozygous mutants were already gained, one T-DNA insertion mutant was named sb3type, another transposon insertion mutant was named cs21type.Comparison of homozygous mutants and wild-type Arabidopsis thaliana showed that there were no significant differences in phenotype.5.The full-length CDS of AtSb3gene was cloned, the overexpression vector pBI121-35S::AtSb3was successfully constructed. Overexpression transgenic lines were obtained by floral dip method of Arabidopsis thaliana. Semi-quantitative RT-PCR anlysis of transgenic lines indictated AtSb3was overexpressed in transgenic plants.