Background: Among all blood group systems and antigens, ABO and D antigens of the Rh blood group system are of primary importance, hence included in routine blood grouping and transfusion. Other blood group systems and antigens that are important in multiparous women, multi-transfused patients and hemolytic disease of newborn are Kell, Rh, Duffy, and Kidd.Aims and Objectives:  To know the pattern of ABO, RhD and other clinically significant major antigens of Rh, Kell, Duffy and Kidd blood group systems as well as evaluation of phenotypes, genotypes and gene frequency of related antigens among blood donors in Gwalior region.Materials and methods: A prospective study was carried out at the Blood Bank from July 2017 to June 2019. During the study period 48500 blood donors were included for ABO and RhD grouping, Out of them 1000 randomly selected donor samples were processed for complete Rh, Kell, Duffy and Kidd blood grouping.Result:  ABO group pattern was; A - 22.56%, B -  36.52%, AB - 9.8% and O -31.12 %, while RhD status was RhD+ 90.99% and RhD- 9.01% . Prevalence of Rh phenotypes was DCCee 43%, DCcee 33%, DCcEe 10%, dccee 6.5%, DccEe 4.5%, Dccee 1%, DCCEe 0.3% and dccEe 0.2%. Kell phenotype was K-k+ 95.5%, K+k+ 4.5%, while K+ k- and K- k- phenotypes were not encountered in this study. Duffy phenotypes were Fya+ Fyb+ 47.5%, Fya+ Fyb- 35%, Fya- Fyb+17% and Fya- Fyb- 0.5%. Kidd phenotype was Jka+ Jkb- 44.5 %, Jka- Jkb- 26.5%, Jka+ Jkb+ 18% and Jka- Jkb+ 11%.Conclusion: Present study is helpful in the transfusion management of multiparous women and multi-transfused patients. Extended blood grouping is the need of future to ensure safe blood transfusion practices.