The aim of the studyis to evaluate the suitability of STRs for molecular characterization and forensic applications in unrelated Brahmins of Rajasthan and Haryana states, India.Materials and MethodsA total of 203 male DNA samples from various districts of Haryana (n=104) and Rajasthan (n=99) were genotyped using the GlobalFiler®PCR Amplification Kit. Allelic frequencies and different forensic parameters like PD, PE, PIC, PM, Ho, He, UHe, and TPI were calculated with different software.ResultsMore than 200 alleles were present in both populations, ranging from 6.0 to 35.2 and SE33 was the most polymorphic marker. The combined power of discrimination was 1. To know the relatedness with other Indian Brahmin populations, the UPGMA dendrogram and principal component analysis plot were visualized to show that both populations are close to each other and in nearby Saraswat Brahmins of Himachal Pradesh. This study showed a genetic relationship and forensic examination in the Haryana and Rajasthan Brahmin populations and various ethno-linguistically diverse populations of India.ConclusionThe results imply that the highly polymorphic 21 autosomal STR loci might be applied for individuals’ forensic identification and parentage testing. This study also suggests that the kit having both autosomal and Y-STR markers is appropriate for a better understanding of the genetic and forensic examination in the Brahmin population of Haryana and Rajasthan.