Cloning And Expression Analysis Of Three Dehydrin Genes In Wheat

Based on the data obtained from the gene expression profiles indrought-stressed roots of common wheat, three drought-induced dehydrin genes werecloned by Reverse Transcription-Polymerase Chain Reaction (RT-PCR), and theirexpression patterns responding to abiotic stresses, such as drought, salt heat and cold.Additionally, the effects of dehydins on the protection of enzyme activities werepreliminary tested. The main results were showed as follows:The cDNA of TaDHN-1is487bp in length and no introns are included, whichencodes112amino acids with a molecular weight of11.5kDa. A typical dehydrinconserved domain which is sorted into Kn subfamily of dehydrins in TaDHN-1. Thecoding region of TaDHN-2is556bp which encodes152amino acidsand there is a109bp intron themolecular weight of TaDHN-2is15.55kDa, Amino acid sequenceanalysis indicates that TaDHN-2belongs toYSKn subfamily of dehydrins. The codingregion of TaDHN-3is589bp which encodes150amino acids and there is a83bpintron. The molecular weight of TaDHN-3is15.3kDa, an typical dehydrin conserveddomain which is sorted into YSKn subfamily of dehydrins is found in it; The structureprediction of the three dehydrin genes revealed that there are no transmembraneregions and signal peptide, and both of them are located in cytoplasm.Expression patterns analysis revealed that all of TaDHN-1, TaDHN-2andTaDHN-3were up-regulated by ABA-induce and drought stress in both Luohan2andChinese Spring. Considering of the two varieties tested, the induced expression levelof the three dehydrin genes was significantly higher in Chinese Spring than Luohan2generally, this may because that Luohan2was more drought-tolerance than ChineseSpring; Never the less, the expression level of TaDHN-2in leaves was higher in Luohan2during drought stress which may indicated a different drought stress respondingmechanism in leaves between TaDHN-2and the other two dehydrin genes. Expressionpattern of the three dehydrin genes was far more different during high-salt stress;Expression level of TaDHN-1and TaDHN-2was up-regulated in root during salt stress, and was higher in Chinese Spring, expression level of TaDHN-2was very highin the root of Luo han2under salt stress but was only induced at0.5h in the root ofChinese Spring. Induced expression of the three dehydrin gene was slower in leavescompared with root, and expression level in Luo han2was significantly higher thanChinese Spring, this result may indicate a different responding mechanism under saltstress between leaves and root. Induce model in the root of wheat was similar betweenTaDHN-1and TaDHN-2under low-temperature stress, but expression level ofTaDHN-1was higher in Luo han2than that in Chinese Spring and TaDHN-2was onthe contrary. Comparatively speaking, TaDHN-3was insensitive for low-temperature,we only observed a higher expression level in Luo han224h after treatment.Expression level in leaves of TaDHN-1was both up-regulated in the two varietiesunder low-temperature stress but higher in Chinese Spring versus Luo han2; Inducedexpression level in leaves of TaDHN-2was significantly higher in Luo han2compared with Chinese Spring, however, no obvious responds was observed forTaDHN-2in leaves under low-temperature stress. What’s more, induced expressionpattern of TaDHN-2was on the contrary between Luo han2and Chinese Springwhich may because of the different vernalization characteristics of the two varieties.Generally, no significantly induced expression in leaves were observed for the threegenes under high-temperature stress, and the up-regulated expression level after24hstress was probablely because of the withered and dehydration of the plant.Expression level in leaves was significantly up-regulated for all of the three dehydringenes in Luo han2under high-temperature stress after12h and this was consistentwith the strong stress resistance ability of Luo han2. There is usually a process ofdehydrin and drying during the development of wheat seed. We examined theexpression pattern of the three dehydrin genes mentioned above and the resultsshowed as follows: expression level of TaDHN-1was down-regulated along with thedevelopment of the seed and was extremely low in the late period. We inferred thatTaDHN-1was not involved in the dehydrin protection process during advanced stateof the seed maturity. Expression level of TaDHN-2was extremely up-regulated in theseed of Chinese Sping after25days after pollination which is a key turning point forthe dehydrin of the seed. We inferred that the up-regulated expression level ofTaDHN-2may related to the launch of dehydrin of the cell, however, no obviouslydifferent expression pattern was observed in the seed of Luo han2which mayindicate that TaDHN-2was not participant in the dehydrin protection of Luo han2. Expression level of TaDHN-3was both up-regulated in Luo han2and Chinese Springduring seed development, and expression level in Luo han2was lower than ChineseSpring. We inferred that TaDHN-3may involved in the dehydrin protection process ofthe cell during seed development. To further analysis the protection effects onenzyme activity of dehydrin, the expressin in vitro and preliminary identificationactivity showed that dehydrin TaDHN-2played some protective role on a-amylaseactivity.