Expression Analysis Of Peanut Root-specific Promoter AhMtan In Tobaccos

Arachis hypogaea L.is one of the world's most important crops,rich in protein and oil.Underground pests,especially alfalfa,are seriously harmful to them,and their roots and pods are foraging,which seriously affects the quality of peanuts.However,common control methods have potential drawbacks for their prevention and control.Therefore,the use of plant genetic engineering methods for green control is extremely important.It is important to look for efficient tissue-specific promoters to drive the expression of foreign target genes in specific tissues.This study is based on the pre-cloned root-specific promoter,further functional analysis and validation,and the use of root-specific promoter to drive the expression of cry8-like gene in tobacco,thus laying a theoretical foundation for the development of anti-tuberculosis crops.The main findings are as follows:(1)Functional analysis of the 1021 bp Ah Nramp1 promoter was performed in the previous clone,and the 5' end deletion analysis was performed on the promoter.Two truncated promoters(796 bp,746 bp)were constructed and fused with the GUSPlus reporter gene to obtain 16H796.And 16H746 two plant expression vectors.A total of 20 kanamycin resistant seedlings have been obtained by genetic transformation of tobacco by Agrobacterium tumefaciens.(2)Further functional analysis of the synthetic promoter SRSP,RT-PCR results showed that the gus gene driven by the SRSP promoter was only transcribed at the root of transgenic tobacco.GUS chemical histological analysis of whole T1 transgenic tobacco was carried out.The synthetic promoter SRSP could drive the gus gene to express specifically in tobacco roots,but no gene expression in leaves and veins.(3)The preliminary functional analysis of the peanut Ah Mtan promoter was carried out.GUS chemical tissue staining was performed on the whole plant transformed with tobaccos.Microscopic observation showed that the promoter could specifically express the reporter gene gus at the root and paraffin section of the root of the transgenic tobaccos.Transverse and longitudinal observations were performed to show that the gus gene was expressed in the vascular column and cortex of the root,and the epidermis was not expressed.The Ah Mtan promot er contains only two root regulatory elements,ROOTMOTIFTAPOX1 and OSE2 ROOTNODULE,which are related to these two components.(4)The cry8-like gene expression vector driven by Ah Nramp1,Ah Mtan1 and SRSP promoters was constructed respectively.The recombinant plasmid was transferred into Agrobacteriumtumefaciens LBA4404,and the tobaccos was genetically transformed by Agrobacterium-mediated method.After PCR identification,6 positive seedlings were obtained.This study provides a basis for subsequent insect resistanceIn this study,the peanut root-specific promoter Ah Nramp1? Ah Mtan and the synthetic promoter SRSP promoter obtained in the early stage of the laboratory were further functionally verified,and the expression of the cry8-like gene with insecticidal activity in tobacco was used in tobaccos,This laid the foundation for obtaining Grub-resistant peanuts.