Molecular Characterization And SNP Markers Of The β-purothionin Gene In Einkorn Wheats

Based on the homologous cloning,forty-three gene sequences encoding purothionin were characterized from the three species or subspecies of einkorn wheats.Genetic variations of purothionin in molecule level were detected,and then a number of SNP loci were detected,and the special SNP markers were developed.Amino acid variations caused of cSNPs were discussed in our study.The following results were obtained.1.The purothionin gene of 43variaties were isolated and sequenced,the result showed that 70 SNPs including 12 indel loci were detected,giving an average SNP frequency of one SNP per 12.23 bases.Twenty-nine SNPs occurred in extron,and the remainings were observed in the two introns.According to these sequences,5 SNP markers were successfully designed,which were used to detect the variations of purothionin genes in 102 einkorn wheat accessions.Based on the 5 detected SNP loci,102 einkom wheat accessions could be divided into 21 haplotypes,among which 11 haplotypes contained a single sample.2.Phylogenetic analysis of purothionin genes from 43 einkorn wheats indicated that they could not cluster according to species,subspecies or geographical sources.The purothionin genes were not species-speciality and were not related to the geographical origin.Additionally,it also indicated that purothionin genes from A genome were more closely related to those from D genome other than B genome.3.Eight out of the 43 gene sequences were assumed to be pseudogenes by the definition of containing in-frame stop codons and small insertions/deletions leading to abridged translational protein sequences.Eight pseudogenes just occurred in T.boeoticum and T. uratu which were the wild types of einkom wheats.SNPs in the extron resulted into 14 amino acid variations among which 2,7 and 5 were observed in signal peptide,mature thionin domain and in carboxy-terminal acidic domain,respectively,indicating that there were more variations in mature thionin domain.Amino acid residues played important roles in the biological activities,such as the 8 Cys,Lys-1,Ser-2,Arg-10,Tyr-13,Arg-17 and Gln-22 loci in the mature thionin domain which were all conserved. The purothionins had similar crystal packing associated to form dimmers,Asn-11 and Asn-14 was important to this process.In the deduced amino acid sequence of pth42, there was a variation of Asn11-Asp11,which could influence packing of the dimmer.