Variability of Ribosomal DNA ITS-2 and Its Utility in Detecting Genetic Relatedness of Pearl Oyster

The objective of this study was to detect interspecific and intraspecific genetic variations of the second internal transcribed spacer of ribosomal DNA (ITS-2), and explore the feasibility of using it as a molecular marker phylogenetic analyses and species identification among pearl oysters. ITS-2 sequences of 6 pearl oysters were amplified via polymerase chain reaction. The amplified DNA fragments were about 500 bp, spanning the partial sequences of 5.8S and 28S rRNA genes. The GC contents of all species used in this study were higher than the AT contents. The variations of sequences involved substitutions as well as insertions/deletions and were mainly concentrated in spacer regions. Sequences of about 30-bp in spacer regions showed no variations among 5 Pincatda species. Intraindividual and intraspecific polymorphisms of ITS-2 sequences were detected in some species; the interspecific variability was significantly larger than the variability within species, and the variability at the genus level was higher than that at the species level. Both neighbor-joining and parsimony analyses of ITS-2 sequences revealed the distinguishable species boundary of 6 pearl oysters, and indicated that P. chemnitzi and P. nigra were the closely related species, as were P. maxima and P. margaritifera. The findings revealed that ITS-2 sequences could be an appropriate tool for phylogenetic study of pearl oysters.