Synthetic virology is an important multidisciplinary scientific field, with emerging applications in biotechnology and medicine, aiming at developing methods to generate and engineer synthetic viruses. Here we demonstrate a full multidisciplinary pipeline for generation and analysis of synthetic RNA viruses and specifically apply it to Dengue virus type 2 (DENV-2). The major steps of the pipeline include comparative genomics of endogenous and synthetic viral strains. In particular, we show that although the synthetic DENV-2 viruses were found to have lower nucleotide variability, their phenotype, as reflected in the study of the AG129 mouse model morbidity, RNA levels, and neutralization antibodies, is similar or even more pathogenic in comparison to the wildtype master strain. These results may suggest that synthetic DENV-2 may enhance virulence if the correct sequence is selected. The approach reported here can be used for understanding the functionality and the fitness effects of any set of mutations in viral RNA. It can be also used for editing RNA viruses for various target applications.