The aim of the present study was to survey mtDNA 16S rRNA sequence variation to assess genetic diversity and genetic structure of the wild tiger prawn in the coastal waters of South China. The samples of P. monodon had been obtained from the coastal waters of South China which included the coastal waters of Sanya(SY), Shenzhen(SZ), Zhanjiang(ZJ), Yangjiang(YJ), and Beihai(BH). 100 individuals of P. monodon were caught from the five putative populations (SY putative population, SZ putative population, YJ putative population, ZJ putative population and BH putative population). The PCR technique was used to amplify the mtDNA 16S rRNA among 100 individuals. The PCR products were purified and sequenced. As a result, 440 bp nucleotide sequences of partial 16S rRNA were obtained (some of the marginal sequences were excluded). 19 haplotypes and 28 variable sites were detected among all partial mitochondrial 16S rRNA sequences of 100 tiger prawns. Nucleotide diversities (π) in SY putative population, SZ putative population, YJ putative population, ZJ putative population and BH putative population were 0.004 35，0.005 86，0.010 50，0.010 81 and 0.011 68 respectively. Haplotype diversities (H) in SY putative population, SZ putative population, YJ putative population, ZJ putative population and BH putative population were 0.6895, 0.5211, 0.5737, 0.6000 and 0.7211 respectively. F_ST values between ZJ and SY，SZ putative populations, between BH and SY，SZ putative populations showed significant genetic differences， and F_ST values among all the other populations showed no strong genetic differences. Significant genetic differentiation among all five putative populations was found by analysis of molecular variance (AMOVA). The NJ tree of the five putative populations showed the SY and SZ putative populations had the closest relation among the five putative populations; the BH and ZJ putative populations had the greatest genetic distance from the SY and SZ putative populations among the five putative populations. Therefore, we conclude that the five putative populations can provide two basic populations for selective breeding of P. monodon; one is the SY and SZ geographic stock, and the other is the ZJ and BH geographic stock.