Mutation breeding has brought significant contributions to the development of high value crops. It steered the first studies to generate plants with desired mutations of genes encoding key enzymes involved in important metabolic pathways. Molecular characterization of 5-methyl tryptophan (5-MT) resistant plants has revealed different base changes in alpha unit of anthranilate synthase (OsASA) gene that can lead to insensitivity to feedback inhibition of anthranilate synthase. The objective of this study was to perform in silico analysis of microarray data from five progressing time points during grain filling of rice. Results showed various differentially expressed genes. Enrichment of these genes revealed their roles in amino acid transportation during grain filling. Surprisingly, among all DEGs, only LOC_Os06g42560, a tryptophan synthase beta chain, was found to be directly related to tryptophan biosynthesis. It might affect amino acid content during grain filling. For physico-chemical analysis, different grain and eating qualities parameters were measured using mutant rice lines. Evaluation results showed that 5MT resistant-lines (5MT R-lines) showed approximately 60% chalkiness after milling although it had 20 times higher tryptophan content measured in μg/100 mg seeds. Taste quality of these 5MT R-lines in general was not affected significantly. However, other parameters such as peak time of viscosity and gelatinization temperature showed different results compared to the wildtype. Mutant lines generated in this study are important resources for high tryptophan content, although they have lower grain quality than the wildtype. They might be useful for developing new high nutrient rice varieties.