Cloning And Function Analysis Of FaMADS1 And FaMADS2 Genes From Strawberry

MADS-box plays an important rool in the development of roots,leaves,flowers and fruits,and the maturation of fruits.MADS is a key gene family that regulates the reproductive growth of plants and the development of vegetative organs.Strawberries are typical non-climacteric fruits,and ethylene-independent regulatory pathways play a major role in strawberry fruit maturation.The MADS-box transcription factor can regulate the maturation process of many fruits.In this study,we cloned two MADS-box genes,which named as FaMADS1 and FaMADS2.The expression patterns of the two MADS-box transcription factors are opposite in strawberry fruit development.It is speculated that the role of strawberry fruit ripening is different.In this paper,the function of FaMADS1 and FaMADS2 genes was identified to analyze the roles of two MADS-box genes in strawberry ripening.This study not only provides in-depth understanding of the role of MADS-box genes in non-climacteric fruit maturation,but also provides theoretical support and gene reserve for the enhancement of strawberry’s resistance to storage and transportation through heritage measures.It has important theoretical significance and practical application value.In this study,we used sweet charlie strawberry fruit as a test material.Two MADS-box transcription factors,FaMADS1 and FaMADS2,were cloned and related bioinformatics analysis was performed.Real-time quantitative PCR was used to analyze the expression level of FaMADS1 and FaMADS2 at different developmental stages and in four different organs of sweet Charlie.Overexpression vectors 35S:FaMADS1 and 35S:FaMADS2 were constructed and genetically transformed into Arabidopsis thaliana and tomato for gene functional validation studies.The main results for this study are as follows:1.Two MADS-box genes,FaMADS1 and FaMADS2,were cloned and obtained from sweet Charlie strawberry fruit 30 days after flowering.The sequencing results showed that the ORF of FaMADS1 consisted of 768 bases and encods a protein of 255 amino acids.The results of protein multiple sequence alignment showed that it has higher homology with tomato SlRIN.The ORF of FaMADS2 consists of 621 bases and encodes a total of 206 amino acids.The results of protein multiple sequence alignment showed that it has high homology with Arabidopsis thaliana AtAP3.2.Protein domain analysis showed that both FaMADS1 and FaMADS2 contain highly conserved MADS-box and K-box domains.3.By analyzing the spatiotemporal expression patterns of four different developmental organs of young leaves,functional leaves,flowers and stems,the relative expression levels of FaMADS1 and FaMADS2 in flowers were the highest.Expression patterns of fruits at different developmental stages of strawberry showed that the expression level of FaMADS1 increased during fruit development and was highest at the ginkgo stage.FaMADS2 expressed at a higher level during fruit development and decreased rapidly during the ginkgo period.The expression patterns of the two genes are opposite during fruit development,and it is speculated that FaMADS1 and FaMADS2 have different functions during fruit development.4.Overexpression vectors 35S:FaMADS1 and 35S:FaMADS2 were successfully constructed.35S:FaMADS1 successfully transformed Arabidopsis thaliana and 5 transgenic plants were obtained.35S:FaMADS1 and 35S:FaMADS2 successfully transformed Micro-Tom tomato.Eight FaMADS1 transgenic tomatoes and 3 FaMADS2 transgenic tomatoes were identified.5.Overexpression of FaMADS1 and FaMADS2 gene in strawberry fruit was induced by injection.FaMADS1 did not change the maturation process of strawberry fruit in the injection area,but FaMADS2 inhibited the maturation process of strawberry fruit in the injection area.It was speculated that FaMADS2 could inhibit fruit ripening during fruit development.The function,while FaMADS1 does not inhibit fruit ripening during fruit development.