Isolation and characterisation of ten microsatellite loci from a Western Australian tree, Banksia sessilis (Proteaceae)
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MICROSATELLITE LETTERS
Isolation and characterisation of ten microsatellite loci from a Western Australian tree, Banksia sessilis (Proteaceae) Heidi Nistelberger • Shelley McArthur David Coates • Margaret Byrne
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Received: 12 December 2014 / Accepted: 16 December 2014 Ó Springer Science+Business Media Dordrecht 2014
Abstract Ten nuclear microsatellite markers were developed for Banksia sessilis, a tree endemic to Western Australia to assess whether patterns of genetic diversity and structure reflect the taxonomic varieties currently described, one of which is a priority species for conservation. One tetranucleotide, three trinucleotide and six dinucleotide repeat loci were tested on 24 individuals from each of two populations. All loci showed independent inheritance and were polymorphic. The number of alleles ranged from two to seven. Observed heterozygosity ranged from 0.083 to 0.750 and expected heterozygosity from 0.080 to 0.718. Keywords Genetic diversity Banksia sessilis Microsatellites Proteaceae Banksia sessilis (Knight) A.R. Mast & K.R. Thiele, commonly known as Parrot Bush, is a small tree endemic to the South Western Australian Floristic Region. It is currently comprised of four taxonomic varieties, var. cordata, var. cygnorum, var. falbellifolia and var. sessilis, based on differences in morphology and distribution. One of the varieties, B. sessilis var. cordata has a limited distribution along the southern, Western Australian coastline, and is considered a Priority Four (rare/near threatened) species of conservation significance by the Western Australian
Electronic supplementary material The online version of this article (doi:10.1007/s12686-014-0409-z) contains supplementary material, which is available to authorized users. H. Nistelberger (&) S. McArthur D. Coates M. Byrne Science and Conservation Division, Department of Parks and Wildlife, Bentley Delivery Centre, Locked Bag 104, Bentley, WA 6983, Australia e-mail: [email protected]
Department of Parks and Wildlife (Smith 2013). This translates to an IUCN classification of ‘Data Deficient’. Microsatellite markers were developed to determine whether patterns of genetic diversity and structure reflect the current taxonomic division. Genomic DNA was extracted from lysed, freeze-dried leaf material (approx. 40 mg) using a 2 % CTAB protocol (Doyle and Doyle 1990) that substitutes diothiothreitol with b-mercaptoethanol and ethanol (95 %) with isopropanol. One individual was sequenced on a 454 shot-gun GS-FLX platform (Roche), located at the Australian Genome Research Facility (Adelaide) to allow isolation of microsatellite sequences according to Gardner et al. (2011). Initially, 35 Primer pairs were designed with PRIMER3 ver. 1.1.1 [see Gardner et al. (2011)] and used to screen six individuals from three populations to test for successful amplification and polymorphism using 3 % agarose gels. Of these 35 pairs, 12 produced clear banding patterns on the gel and these were then used to amplify DNA from a population of indivi
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