The Mechanism Of BpNAC2 Transcription Factor In Regulating Salt Tolerance Of Betula Platyphylla

NAC transcription factors are a class of transcription factors that are unique to plants.They have multiple biological functions and play an important role in responding to abiotic stress.In this study,a differentially expressed gene BpNAC2 was selected from the regulatory network of Betula platyphylla in response to osmotic stress and study it in detail.The full length of BpNAC2 gene is 891 bp,encoding 296 amino acids.Analysis of its amino acids showed that it has a typical NAC conserved domain.Multiple sequence alignment showed that BpNAC2 had the highest homology with ANAC2.The pBI121-BpNAC2-eGFP overexpression vector was constructed.Transient transfection Nicotiana benthamiana and the results of laser confocal observation showed that BpNAC2 was a nuclear protein.The expression of BpNAC2 gene was highly specific in different tissues,with the lowest expression level in roots and the highest expression level in terminal buds.BpNAC2 can respond to NaCl and ABA stress induction,and has significant changes in roots.According to tissue-specific analysis,BpNAC2 expression level is the lowest in roots under normal growth environment,but BpNAC2 is significantly induced in roots under NaCl and ABA stress.These results suggest that BpNAC2 may respond to salt stress by participating in the regulation of root growth and development..Stable overexpressed transgenic plants(OE)of birch were obtained by agrobacteriummediated leaf disk transformation.Three lines with high expression(OE-1,OE-5 and OE-10)were selected for study.Wild-type plants and overexpressed plants were treated with salt stress,and their phenotypes were photographed and statistically analyzed.It was found that the overexpressed lines had stronger salt tolerance,and the roots of the overexpressed lines were developed and the taproots were longer under normal and salt stress,suggesting that BpNAC2 might be involved in the root growth and development of birch.Measurement of relevant physiological indicators: The results of DAB and NBT staining on birch leaves showed that the accumulation of reactive oxygen species(ROS)in overexpressed plants was significantly decreased,POD and SOD activities of overexpressed plants were significantly increased,malondialdehyde(MDA)content and relative electrical conductivity of overexpressed plants were significantly decreased compared with control.These results showed that BpNAC2 overexpression could improve ROS scavenging ability,and BpNAC2 overexpression could regulate stomatal opening to reduce water loss in response to salt stress,thus significantly improving salt tolerance of birch.The results obtained by RNA-seq and ChIP-seq were combined to analyze the downstream regulation genes of BpNAC2,the results show that the up-regulated differential genes include genes related to peroxidase,auxin-responsive protein and potassium ion transporter,and the New PLACE database was used to analyze the promoter regions of related genes.It was found that G-box,"GGGCCC" and CCAAT box were identified in ChIP-seq,and these elements appear more frequently.It is suggested that BpNAC2 transcription factor may directly regulate these genes to promote plant root growth and development,improve ROS scavenging ability,and regulate stomatal opening in response to salt stress.At the same time,the interaction between BpNAC2 and G-box and "GGGCCC" elements was verified by EMSA.The results showed that the BpNAC2 transcription factor can specifically recognize G-box,"GGGCCC",and CCAAT box regulating the expression of downstream genes and improve the salt tolerance of birch.