DNA marker technologies for biodiversity study in shrimp

Four DNA extraction methods were tested for isolation of high molecular weight genomic DNA from shrimp. The proteinase K method modified from McCouch et al. (1988) gave the higher quality and quantity than others. Furthermore, genomic DNA also can be extracted from one month old of formaldehyde or ethanol fixed tissues using this extration method. However, formaldehyde fixed tissue gave very low DNA yield. Total genomic DNA from shrimps were used for genetic diversity evaluation using three DNA marker technologies, mitochondrial-specific markers, random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP). For mitochondrial-specific markers, eight shrimp species were examined using specific primers for 12s and 16s rDNA. Two monomorphic products of 415 bp and 520 bp fragments were obtained from 12s and 16s, respectively. After digestion of those monomorphic fragments with nine restriction enzymes, gave the total of 77 scorable loci. Percentage of polymorphic loci and the Nei and Li similarity coefficient were ranging from 33.33-88.74 percent and 0.1026-0.6667, respectively. Twelve shrimp species analyzed by three RAPD primers (OPAE06, OPAE07, OPAE09) gave the total of 51 scorble loci. Percentage of polymorphic loci and the the Nei and Li similarity coefficient were ranging from 48.34-100 percent and 0.0000-0.5263, respectively. For the AFLP analysis, twelve shrimp species analyzed by two primer combinations (ER-GTC, MS-ACC and ER-GTC, MS-ACT) gave 150 scorable loci ranging from 200 bp to 726 bp in size. Percentage of polymorphic loci and the Nei and Li similarity coefficient were ranging from 33.33-100 percent and 0.000-0.6667, respectively. Genetic relationship among shrimp species was clearly shown by the dendrogram generated from mitochondrial-specific markers analysis. The species belong to the same genera were clustered to the same branch, and the grouping made using this technique is similar to the morphological classification. On the other hand, RAPD and AFLP gave too many genetics details causing ambiguous result for classification analysis among shrimp species, especially the species which closely related on the morphology to one another. However, these techniques could be used to evaluate genetic variation of within species.