Nine microsatellite loci were identified in the western rock lobster (Panulirus cygnus) using two different methods. The first method involved the screening of a small, fragment, partial genomic library with a radioactive (CA) 6 probe. The second method, was based upon an enrichment method and used biotinylated, tetranucleotide microsatellite oligonucleotide capture probes. The nine loci described are all very polymorphic, with 11 to 34 alleles observed for each locus and heterozygosities ranging from 0.58 to 0.86. These microsatellite loci will be useful in analysing both the population structure and the mating systems used by this species and will add important information for the management of the wild stocks of this economically important species.Keywords Panulirus cygnus Á Microsatellite Á SSR The western rock lobster (Panulirus cygnus) is Western Australia's most important single species fishery with annual harvests over the last decade ranging between approximately 9 and 14.5 thousand tonnes per annum. The elucidation and understanding of population and mating structures for this species will allow the information obtained to be incorporated into an improved management strategy for the brood stock. In order to understand the population structure and mating systems of the western rock lobster, polymorphic, microsatellite genetic markers have been isolated and characterised. Two strategies were used to develop these microsatellites, the first consisting of the construction of a Hae III small fragment library which was screened with a (CA)n probe and the second library was made using an enrichment technique followed by screening with tetranucleotide microsatellite motifs. Nine microsatellite loci have been fully characterised and these are described in this report.A DNA genomic library was made using Qiagen Tissue Kit (Qiagen) to isolate DNA extracted from tail muscle tissue. The DNA was digested with HaeIII restriction endonuclease (Promega, CA, USA) followed by excision of the 100-500 bp region after agarose (1%) gel electrophoresis. Fragments were ligated into Sma I (Promega) digested, dephosphorylased pUC18 plasmid vector and transformed into competent cells (DH5a Invitrogen). Colonies were transferred to Hybond N? membranes and screened with radiolabelled (CA)10 probe. Thirty clones were identified after the first round of screening. Following successive screening rounds to further isolate the clones, twenty of these were sequenced. Plasmid DNA was isolated from the positive clones and sequenced using cyclesequencing combined with IRD 800 labelled primers on a Li-Cor gene sequencer. Primers were designed using Mac Vector (Eastman-Kodak, USA) or PRIMER 3 software (Rozen and Skaletsky 2000). Many of the loci identified contained insufficient flanking DNA sequences or produced microsatellites containing few repeat units. Three loci (WRL 1, WRL 2 and WRL 3) identified by using this method were deemed to be useful.A second library enriched for tetranucleotide microsatellites was prepared using genomic DNA. Br...
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