Nucleotide Polymorphisms At The Drought Candidate Gene Loci Of Arabidopsis Thaliana And Their Association Study With Drought Index And Functional Study Of ATHK1

The levels of drought tolerance and nucleotide polymorphism at transcription factor CBF4, ATHK1 and key enzyme regulating abscisic acid (ABA) synthesis NCED3 locus in a worldwide sample of 32 core accessions of Arabidopsis thaliana were examined. The results showed that different accessions had quite difference in adaptation to drought stress. Thirty-two core accessions of Arabidopsis thaliana were classified in four clusters by synthetical analysis of drought tolerance.The frequency of nucleotide polymorphism at the CBF4 locus, including single nucleotide polymorphism ( SNP ) and insertion/deletion( Indel ), was high, on average 1 SNP per 35.8bp and 1 Indel per 143bp. In coding region, SNP frequency was 1 SNP per 96.4bp, one nonsynonymous mutation was detected, which was the 205th site amino acid variation: gly val caused by the 1034th site (corresponding to 19696 site nucleotide of GenBank No.ABO 15478 as 1) nucleotide variation: G T. Nucleotide polymorphism in non-coding region was three times higher than that in coding region. No significance (NS) Tajima'D test in all region regions indicated that the neutral mutation hypothesis could explain the nucleotide polymorphism in CBF4 gene region. The higher polymorphism was the result of purification selection. This result might also indicate a recent relaxation of the selection pressures on non-coding region of CBF4 gene. Association study showed that nucleotide variation of 5'flanking sequence of CBF4 was associated with the synthetical drought tolerance, but the amino acid variation of CBF4 was not associated with the synthetical drought tolerance.The frequency of nucleotide polymorphism at the ATHKl locus was on average 1 SNP per 142 bp and 1 Indel per 1705 bp. SNP frequency was 1 SNP per 190.6 bp in coding region and 1 SNP per 87.9 bp in non-coding region, SNP in non-coding region was 1.17 times higher than that in coding region. Significant Tajima'D test hi whole region and coding region indicated that the neutral mutation hypothesis could not explain the nucleotide polymorphism in ATHKl gene region, the higher polymorphism was the result of natural selection. The ratio Ka/Ks, being 0.727, indicated that ATHKl was the middle evolution-speed gene. Association study showed that nucleotide variation of 5'flankingsequence of ATHK1 was not associated with the synthetical drought tolerance, but the the amino acid variation of ATHKl of 257av and 266av, which was 1198 site Ser stop code, was associated with their lower synthetical drought tolerance.The frequency of nucleotide polymorphism at the NCED3 locus was on average 1 SNP per 87.4 bp and 1 Indel per 502 bp. SNP frequency was 1 SNP per 94.6 bp in coding region and 1 SNP per 51 bp in non-coding region. SNP in non-coding region was 0.85 times higher than that in coding region. No significance Tajima'D test in whole region and coding region indicated that the neutral mutation hypothesis could explain the nucleotide polymorphism in NCED3 gene region, The higher polymorphism was the result of purification selection. But significant TajimaT) test in 5 'flanking sequence indicated that nucleotide variation of 5 'flanking sequence was not explained by the neutral mutation hypothesis but by balancing selection. The ratio Ka/Ks, being 0.2727, indicated that NCED3 was the highly conservative gene. Association study showed that the 33th site A C and 170th site A C nucleotide variation of 5'flanking sequence of NCED3 of 157av was associated with its higher ABA content; 87th site C T of NCED3 of 257av was associated with its the lower synthetical drought tolerance. The amino acid variation of NCED3 of 163av, which was 46th site Cys Arg, was associated with its higher ABA content. The amino acid variation of NCED3 was not associated with their synthetical drought tolerance.Differences in physiology and gene expression between ATHK1 knock-out mutantcaused by T-DNA insertion and wild type (WT) of Wassilewskija (WS) accession of Arabidopsis thaliana were analysed. Water loss ratio of detached leaf of ATHKl -mutant was...