五莲杨PwuWRKY基因组织特异性表达分析

WRKY transcription factors are usually involved in plant defense responses.However,the role of the WRKY family in the defense of Populus wulianensis is limited.[Objective]To study the new important stress response genes in Populus,providing theoretical basis for revealing the molecular mechanism of poplar resistance to adverse conditions and breeding new resistant poplar varieties.[Methods]Based on the WRKY transcription factors in the response to drought-rehydration stress of P.wulianensis,five genes(including Potri.001G3524,Potri.002G193000,Potri.010G147700,Potri.013G153400 and Potri.008G103300)were screened from the P.trichocarpa genome using BLAST.Using these gene sequences as references and P.wulianensis cDNA as template,five genes(PwuWRKY28-1,PwuWRKY23-5,PwuWRKY48-9,PwuWRKY33-11,PwuWRKY48-20)were cloned.Utilize bioinformatics to analyze the structural features and phylogenetic relationships of PwuWRKY protein and its homologous proteins.Analyze the subcellular localization of PwuWRKY protein in tobacco protoplasts by transformation.Use real-time fluorescence quantitative PCR(qRT-PCR)technology to analyze the tissue-specificity and expression pattern of PwuWRKY gene under different stress conditions.[Result]In this study,evolutionary analysis indicated that PwuWRKY33-11 belongs to the I group of the WRKY family,while the other proteins belong to the IIc subfamily.All five WRKY transcription factors localize to the plant cell nucleus,and except for PwuWRKY48-20,which lacks transcriptional activation activity,the other proteins show transcriptional activation.The tissue-specific expression of five WRKY transcription factors in P.wulianensis was studied in response to abiotic stresses such as drought and rehydration,high salinity,high temperature and low temperature.The results showed that the expression of different members of the WRKY family had obvious tissue specificity.PwuWRKY23-5,PwuWRKY48-9 and PwuWRKY48-20 were mainly expressed in the leaves of P.wulianensis,and the other genes were expressed at higher levels in the roots.PwuWRKY28-1 responded to the above abiotic stresses,and the gene was rapidly up-regulated and then down-regulated.The expression of PwuWRKY48-9 and PwuWRKY48-20 was consistent with that of PwuWRKY28-1 in response to high salt treatment,and the expression trend of these two genes was almost the same in response to low temperature stress,indicating that there was functional redundancy in the response of these two genes to low temperature stress.The above five genes were all treated in response to high temperature stress.The results showed that the expression of WRKY transcription factors was affected by stress,but different WRKY family members responded differently to stress.[Conclusion]The results of this study showed that the expression of WRKY family members in P.wulianensis has tissue specificity,and different members have different responses to non-biotic stress.This expression pattern may be adapted to the stress response of P.wulianensis in different environments.The results of this study lay a foundation for further revealing the mechanism of WRKY protein action and aim to provide a basis for genetic improvement of P.wulianensis.