Exogenous ABA Regulates The Expression Of BZIPs Transcription Factors In Winter Wheat At Low Temperature

Winter wheat(Triticum aestivum L.)Dongnongdongmai1(Dn1)and Dongnongdongmai 2(Dn2)are strong cold-resistant varieties that can safely overwinter in Heilongjiang Province.Has important practical significance.Jimai 22(J22)has strong cold resistance,but it cannot be wintered in Heilongjiang Province.Abscisic acid(ABA)plays an important role in the process of abiotic stress in plants.In this study,we used these three winter wheat as experimental materials to select three transcription factor genes(TabZIP2,TabZIPD and TabZIP-like)in the bZIP family related to stress resistance,and naturally cool in the field(5?,0?,-10?,-25?),explore whether its transcription level expression is regulated by ABA and low temperature.By quantitative method,the relative expression levels of three transcription factor genes at low temperature were determined,and the transcription factors with large ABA correlation change at low temperature were screened and bioinformatics prediction analysis was carried out,and the overexpression of TabZIP2 was cloned and constructed.The vector was transiently expressed in tobacco leaves to analyze the expression of TabZIP2,which complemented the mechanism of transcription factors regulating the cold resistance of winter wheat.The research results are as follows:(1)Sampling at 5?,0?,-10?,-25?(natural cooling in Daejeon),and performing quantitative RT-PCR on three transcription factors(TabZIP2,TabZIPD,and TabZIP-like),the results showed: At low temperature,the expression levels of the three transcription factors TabZIPs in Dn1 bifurcation reached the highest at-10?,indicating that the temperature of-10? is the key temperature for the regulation of cold stress-related transcription factors in Dn1.After exogenous ABA treatment,the expression levels of three transcription factor genes TabZIPs reached the highest at-10?.Before and after ABA treatment in Dn1 leaves,the expression levels of the three transcription factor genes TabZIPs were highest at 0?,indicating that the leaves responded to low temperature earlier than the tiller.In addition to TabZIP-like,the other two transcription factors of Dn2 were the highest at-10?.After ABA treatment,except for TabZIPD,the other two transcription factors were the highest at-25?.TabZIP2 is more sensitive to cold stress after ABA treatment.The expression of Dn2 leaves was the highest at 0? after ABA treatment,which may be due to the early initiation of ABA on the cold resistance mechanism of three transcription factors in Dn2 leaves.The variation of J22 bifurcation and leaf expression was relatively stable,but the leaves were more resistant to cold than the subdivision.(2)TabZIP2 bioinformatics analysis: The full-length cDNA of TabZIP2 gene is 1311 bp.It is predicted by NCBI that TabZIP2 contains a complete open reading frame of 759 bp,which belongs to mixed type,unstable and hydrophilic protein.By predicting the secondary structure of wheat TabZIP2 protein,it was found that ?-helix and random coil accounted for 92.07% of the protein structure.It can be seen from the phylogenetic tree that TabZIP2 is closely related to Brachypodium serrata and rice,that is,the bZIP clustering relationship of these monocotyledons is relatively close.As can be seen from the multiple sequence alignment,the protein has a conserved basic leucine zipper domain.Subcellular localization indicates that the protein is located in the nucleus.(3)The TabZIP2 gene in Dn1 was cloned with a gene size of 759 bp,and the overexpression vector pCMBIA2300u-TabZIP2 of TabZIP2 was successfully constructed.The tobacco leaves were transformed by Agrobacterium injection infiltration method.The results of qPCR showed that the expression of TabZIP2 increased after low temperature treatment,indicating that the expression of this gene enabled tobacco to better adapt to low temperature stress.In other words,the TabZIP2 gene plays an active role in the adaptation of Dn1 to low temperature stress.