The invention relates to a gene combination for producing betaine by expression in rice seed endosperm and an application of thereof, the gene combination comprises a meloS gene, a BvDODA1S gene and aBvCYP76AD1S gene, and further, a nucleotide sequence of the meloS gene is shown as SEQ ID NO.1; the nucleotide sequence of the BvDODA1S gene is as shown in SEQ ID NO.2, and the nucleotide sequence ofthe BvCYP76AD1S gene is as shown in SEQ ID NO. 3, the ORF of the three genes is connected between a rice endosperm specific globulin gene promoter and a terminator to construct a gene box, the gene box is inserted in a plant binary vector, and the betaine-rich rice is obtained, and the obtained rice seeds can be directly used for eating, can also be used as the betaine production raw material seeds, and provide important guiding significance and production application value for the breeding of novel functional crops.