Mt DNA analyses have been shed light on the biogeography of the Asian cavity nesting honey bee Apis cerana Fabricius, 1793. Variations in the CO-I and CO-II regions of mtDNA of the Indian Honeybee Apis cerana indica (Farb.) in Tamil Nadu is studied by using sequence analysis of intergenic region between CO.I and CO.II of mitochondrial DNA. Samples of A. c. indica were collected from 7 locations covering ecological regions like plains and hills. The mitochondrial DNA fragment containing a non-coding region was amplified and sequenced from 594 to 744bps. The haplotype diversity (0.905) and nucleotide diversity (0.16508). Intergenic region between Cyto. I and Cyto. II was sequenced from seven samples. The length of 649 base pairs sequenced in the sample of Kodikanal; 658 bps of Mudhumalai; 653 bps of Mayiladuthurai; 650 bps of Parambakudi; 594 bps of Pichavaram; 746 bps of Udagai; 744 bps of Perambalur. The percentage of A+T is higher in Mayiladuthurai (86.06%), lower in Udagai (72.39%) and G+C higher in sample of Udagai (27.61%) where as lower in sample of Mayiladuthurai (13.94%) among the mtDNA sequences in this study.