Functional Verification Of The LAZY1 Gene In Brassica Napus

Rapeseed is an important oil crop in my country,and edible rapeseed oil accounts for more than 50% of the domestic supply of edible vegetable oil,which is closely related to the lives of people across the country.The reasonable plant type can increase the leaf area index to a large extent,so that the photosynthetic efficiency of the group can be improved to a certain extent.It is of great practical significance for improving the yield of rapeseed,and the branching angle is an important aspect of the rapeseed plant structure..Therefore,it is of great significance to study the genes related to the branching angle of rapeseed and their mechanism of action.In the early stage of this research group,QTL mapping and genome-wide association analysis were used to find the candidate gene Bna C03g06250 D on the C03 chromosome related to the formation of rape branching angle.Some studies have shown that the LAZY1 gene regulates the axillary bud geomorphism by changing the distribution of auxin.Through sequence alignment in the NCBI database,it was found that there are 4 copies of the LAZY1 gene in Brassica napus.In addition to the candidate genes Bna C03g06250 D,there are Bna A03g04710 D,Bna A10g19550 D and Bna C09g54600 D.The research team successfully cloned 4 copies of the CDS sequence in the early stage.The study found that the LAZY1 gene was mainly expressed in the main stem and lateral branches during the early flowering period in Brassica napus.The N-terminal of the LAZY1 gene CDS was located in the cell by subcellular localization.Nuclearly,the C-terminus of CDS is located on the cell membrane.This study mainly carried out the complementary experiment of Arabidopsis mutants;positive identification of transgenic rape;the determination of the auxin content of each main part of the extreme material;at the same time,the core region of the upstream promoter was determined,and the downstream interactive proteins Screened.In this article,the vectors for constructing the Bna A03g04710 D and Bna A10g19550 D genes are collectively named LAZY1-1 and LAZY1-2.The main conclusions are as follows:1.Complementary experiment analysis found that the branching angle of the positive transgenic plants with HY002(compact)Bna A10g19550 D as the gene source changed from greater than or equal to 95°to 42°and grew upright;However,the branching angle of the positive transgenic plants with 03za-1(loose)Bna A10g19550 D as the gene source was also significantly reduced,and some of them had creeping growth.This shows that the LAZY1 gene can indeed regulate the branching angle.2.The positive identification of the transgenic rapeseed found that the LAZY1 gene transgenic plants had a branching angle of 40°,the control ZS11 had a branching angle of 32°,and the branching angle was about 10°larger.The expression level of LAZY1 gene in overexpressed plants was several tens times higher than that of the control.3.Godbole et al inferred that the action site of the LAZY1 gene is between the balance stone sensing gravity and the polar transport of auxin.Therefore,this experiment designed an experiment to determine the auxin content of each part of the extreme material.Through experiments,it was found that the content of LAZY1 gene was positively correlated with the distribution of auxin in the main buds of extreme materials.The auxin content of other parts varies,but in general,almost all materials have more content in the middle and upper branches,which is basically the same as the most gene expression in the middle and upper parts of the early flowering period in the quantitative research of the research group.This speculated that the distribution of auxin is positively correlated with gene expression.4.Through transient expression of tobacco by truncating the 5’end of the promoter,it was found that the core region of the Bna A03g04710D-1500 promoter is located at-130～0bp(with ATG A as +1bp),and the core region of the Bna A10g19550D-1500 promoter is located at-248～-109 bp.The cis-acting element prediction of the 1500 bp promoter core region of Bna A03g04710 D and Bna A10g19550 D was found to contain MRE elements together,and it was speculated that the core promoter element of the LAZY1 gene was MRE(MYB binding site participates in light reaction).It is speculated that the expression of the LAZY1 gene is extremely closely related to the light response,and light will affect the expression of the LAZY1 gene.5.The LAZY1 protein was screened for interacting proteins,and two positive clones were screened.One is Bna Cnng05760 D,its molecular function is responsible for anion transmembrane transport,and the other is Bna C03g75880 D.This gene is a ubiquitin-like domain.Family,responsible for RNA processing.