Mungbean weevil is a type of coleopterans and is one of the main causes of damage to the quantity and quality of mungbean seeds in storage. Weevil resistance of mungbean is regulated by defensin gene. The role of defensin in weevil resistance of mungbean is that causes the inhibition of amylase activity in the digestive tract of weevils. Research on the defensin is the basis for creating transgenic mungbean varieties that are resistant to this type of weevil. In this the works, the authors present the results cloning and determine the nucleotide sequence of defensin1 cDNA from mungbean cultivar. DEF1 cDNA sequences isolated from VN6 variety takes size 0.22 kb, including 222 nucleotides and encodes a respective protein with 73 amino acids. The genetic diversity of VN6 variety and five mungbean varieties from Genbank have been analyzed and evaluated based on the analysis of the nucleotide sequence and amino acid sequence of the DEF1 gene. Genetic distances among six mungbean varieties based on the nucleotide sequence and based on the amino acid sequence of the DEF1 gene is 9.8 percent and 19 percent, respectively. Defensinl cDNA sequence be used to create of vector aims improve ability resistance to weevils in mungbean plants.