Functional Study Of Anthocyanin Synthesis Related Gene FeF3’H1 In Buckwheat

Buckwheat（Fagopyrum esculentum Moench）belongs to Fagopyrum Mill of Polygonaceae,which is one of the main cultivated species of buckwheat.It is a medicinal and edible homologous crop rich in bioactive flavonoids such as rutin,quercetin and anthocyanin,and has high practical value in human nutrition and medicine.In recent years,with the development of social economy and the improvement of human living standards,the demand for functional foods is expanding day by day,and it is of great significance to cultivate new varieties of buckwheat with high anthocyanin content.In this paper,based on the previous transcriptomics analysis of our research group,the FeF3’H1 gene was cloned from buckwheat variety"HHTQ".Through gene overexpression and gene editing technology,the molecular regulation mechanism of FeF3’H1 gene in the process of anthocyanin synthesis in buckwheat was analyzed.The main results are as follows:1.Taking"BHTQ"buckwheat as the experimental material,the inflorescence of buckwheat was infected by Agrobacterium tumefaciens GV3101,and the inflorescence infection transformation method was successfully realized in buckwheat.In addition,it was found that the germination rate of transformed buckwheat was significantly reduced,which was only about 50%of the control buckwheat.When the cell density of Agrobacterium OD₆₀₀ was 0.6,the maximum transformation efficiency was 12.42%,and the transformation system of buckwheat inflorescence infection was successfully constructed.2.By constructing the overexpression vector PHZM69::FeF3’H1 and the gene editing vector pDIRECT＿23D::FeF3’H1 of buckwheat,the expression analysis of transgenic buckwheat progeny showed that overexpression of FeF3’H1 gene could not directly lead to significant changes in anthocyanin content,total phenols and total flavonoids,and could not change the phenotype of buckwheat.Gene editing will significantly reduce the expression of FeF3’H1 gene,down-regulate the expression of DFR and ANS genes,reduce the synthesis of secondary metabolites such as anthocyanins,and finally change the phenotype of buckwheat.The degree of change of buckwheat depends on the degree of gene editing.3.By analyzing the promoter sequences of"HHTQ"and"BHTQ"buckwheat FeF3’H1 gene,it was found that the two buckwheat promoters have the same sequence,and both of them contain related cis-acting elements such as MYB,MYC,light response,plant hormones and abiotic stress.GUS staining showed that the promoter activity of FeF3’H1 gene in two buckwheat plants was very strong.This shows that FeF3’H1 gene plays an important role in the process of anthocyanin synthesis in buckwheat.This shows that FeF3’H1 gene plays an important role in the process of anthocyanin synthesis in buckwheat.