Characterization Of Sucrose Non-fermenting 1-related Protein Kinase 2 Gene TaSnRK2.9 In Wheat (Triticum Aestivum L.)

Sucrose non-fermenting 1-related protein kinase 2(SnRK2),a plant specific protein kinase,is a major hub of abiotic stress signaling,involved in response to external stimuli,plant growth and development,and nutrient utilization.In present study,Ta SnRK2.9s were isolated from wheat(Triticum aestivum L.).The expression patterns of TaSnRK2.9 in wheat seedlings treated with ABA and multiple abiotic stresses were investigated by qRTPCR.Overexpressing TaSnRK2.9 rice lines were used to characterize the gene functions at seedling and mature stage.Based on the SNPs(single nucleotide polymorphisms)detected in the TaSnRK2.9-5A sequences,CAPS(cleaved amplified polymorphic sequence)markers and high throughput gel-free KASP(kompetitive allele specific PCR)markers were developed to identify the haplotypes of the target gene.Association analysis was implemented to investigate the correlations between TaSnRK2.9-5A haplotypes and agronomic traits.The main results of present study are as follows.i)Expression pattern analysis in wheat revealed that TaSnRK2.9 was involved in response to PEG,NaCl,low temperature and ABA treatments.ii)Overexpression of TaSnRK2.9 in rice resulted in enhanced root growth at seedling stage and more grains per spike in rice and also resulted in enhanced tolerance to drought stress,showing more stability in cell membrane and lower chlorophyll a/b.iii)Three copies of TaSnRK2.9 were isolated and designated as TaSnRK2.9-5A TaSnRK2.9-5B and TaSnRK2.9-5D according to their chromosomal origin.The fragments sizes of TaSnRK2.9-5A,TaSnRK2.9-5B,and TaSnRK2.9-5D-PRO were 2180 bp,2137 bp and 2200 bp,respectively.Gene structure analysis showed that TaSnRK2.9 was consisted of nine exons and eight introns.iv)DNA polymorphism analysis results showed that there was no polymorphism in TaSnRK2.9-5B and TaSnRK2.9-5D-PRO.Three SNPs were identified in TaSnRK2.9-5A,i.e.at 308 bp,698 bp and 1700 bp,forming four haplotypes which were identified in Chinese,Pakistani and European wheat accessions.Based on the SNP at 308 bp and 1700 bp,two CAPS markers were developed,validated and successfully converted in to KASP markers.v)Association analysis between TaSnRK2.9-5A haplotypes and agronomic traits demonstrated that Hap-5A-1/2 were associated with higher thousand kernel weight while Hap-5A-4 was associated with more grains per spike.vi)Geographic distribution and allelic frequency showed that favored haplotypes were positively selected in Chinese(Hap-5A-1/2),Pakistani(Hap-5A-1),Eastern Europe(Hap-5A-1),and Western Europe(Hap-5A-4)wheat breeding.The results obtained in this study demonstrated that TaSnRK2.9 has potential for utilization in breeding to improve root traits,drought tolerance,and grains per spike in crop plants.The developed molecular markers can be instrumental in marker assisted selection for genetic improvement and germplasm enhancement in wheat.