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correlograms were applied to the entire dataset as well as to subsets of data (partitioned
according to model based-clustering) using the multivariate, multilocus approach of
Smouse and Peakall (1999) implemented in the program GENALEX 6.3. This method
combines the information generated from multiple genetic markers to strengthen the
spatial signa) by reducing stochastic (allele-to-allele and Iocus-to-locus) noise.
Individuai pairwise genetic distances are used to estimate the autocorrelation coefficient
r, which measures the genetic similarity between pairs of individuals whose geographic
separation falls within a specified distance class. The number and size of distance
classes were set to compare similar sample size within each class. The significance of
positive autocorrelation was determined using both a permutation test (1,000 random
permutations, 95% confidence interval) and bootstrap (1 ,000 reps, 95% confidence
interval) estimates of r. Significant spatial genetic structure was inferred either if the
calculated r value fell outside this confidence interval and ifthe bootstrap confidence

interval did not include r = Ofor a given distance class.

Mitochondrial DNA analysis

The 12S and 16S genes were sequenced for 174 individuals, whereas the COI gene was
sequenced for 110 individuals, being 64 COI sequences available on GenBank
(accession # AY996038-AY996043 ; GQ469872-GQ469874; GQ469876) (Table 1).
Universal primers were used for COI (Folmer et al. 1994), 12S and 16S (Nakano and
Ozawa 2004).

PCR amplification was performed as described in Espinosa and Ozawa (2006)
with the following annealing temperatures: COI, 54 °C; 12S and 16S, 49 °C. PCR
products were purified with ExoSAP-IT (USB Corporation) and were sequenced using

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Gìan Luca Dedola: ANALISI DELLA VARIABILITÀ GENETICA DI PATELLA FéRRUGINEA, PATEL.LA ULl'SSIPONENSIS (MOLLUSCA: GASTROPODA) E PINNA
NOBILIS (MOLLUSCA: BIVALVIA): lLCONTRJBUTO DEl DATI MOLECOLARI ALLA CONSERVAZIONE DI SPECIE MINACCIATE · Tesi di douorato in Scienze
della natura e delle sue risorse • indiri7.zo BK>logi.a Ambientale, Università degli Studi di Sassarl