Recently, genomics has gained a lot of attention in the field of bio-therapeutics, vaccines, and gene therapy. We have developed a robust pipeline for establishing the mRNA vaccine portfolio to address the current pandemic and endemic of COVID-19, infectious diseases, and cancer. We have designed and assembled approximately 4000 base pair DNA encoding the spike protein of the SARS-CoV-2 Omicron variant (B.1.1.529) using overlapping oligo-based PCR assembly. Further, we cloned this fragment into a self-amplifying mRNA platform and prepared the messenger RNA (mRNA). The integrity and sequence were confirmed through multiple orthogonal techniques such as capillary electrophoresis, Sanger DNA sequencing, and Next-generation sequencing for mRNA. The mRNA was transfected into HEK 293T cells and a significant expression of the spike protein was monitored by FACS. Herein, we propose a robust and rapid pipeline for the development of mRNA for vaccines starting from sequence analysis to identifying the lead candidate within 4–5 weeks.