The analysis of 15 autosomal STR locus (TH01, D3S1358, vWA, D21S11, TPOX, D7S820, D19S433, D5S818, D2S1338, D16S539, CSF1PO, D13S317, FGA, D18S51, D8S1179) was done in 582 healthy unrelated individuals (Male-366, Female-216) originating from the various geographical regions of Madhya Pradesh, India. All locus fall under Hardy–Weinberg equilibrium except TPOX. These STR loci were highly informative and discriminating with combined power of discrimination (CPD) >0.99999. Locus wise allele frequencies of the studied population were compared with the other published populations. Also the Clustering pattern and genetic distance of studied populations is compared and presented with various populations. The studied population showed the genetic proximity with geographically close populations of India and significant genetic variation with distant populations which is also evident by clustering pattern of the NJ tree and the PCA plot.