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Fish-R1 and Fish-F1 (Ward et al., 2005), and a 561-bp region of the 16S ribosomal RNA gene was amplified using
primers 16Sar and 16Sbr (Palumbi et al., 1991). All PCRs were performed in 25 µL volumes containing 1 ×
Incomplete NH4 Reaction buffer, 2 mM MgCl2, 0.2 mM dNTP, 0.5 U DFS-Taq DNA polymerase (Bioron GmbH,
Germany), 1 µM of each primer, 80–100 ng of template DNA.
FIGURE 1. Kyphosus vaigiensis (Quoy & Gaimard, 1825) recorded off Favignana Island (37°55’34”N, 12°19’16”E; Egadi
Islands Marine Protected Area, Italy). A: lateral view; B: head; C: first gill arch.
Cycling conditions for PCR amplifications consisted of an initial 95°C denaturation step for 5 minutes
followed by 35 cycles of 60 sec at 95°C, 60 sec at 48°C, and 60 sec at 72°C, with a final extension at 72°C for 8
min and a final cooling at 4°C. The resulting amplified DNA fragments were purified with the QIAquick PCR
Purification Kit (Qiagen), and were sequenced in forward direction, with an Applied Biosystems (ABI) 3730xl
DNA analyzer and the sequences were deposited in GenBank with the Accession numbers: KR013046 and
KR013047. The mitochondrial sequences of the specimen were compared with sequences of congeneric species
deposited in the Barcode of Life Data Systems (BOLD, http://www.barcodinglife.org) (Ratnasingham & Hebert,
2007) and GenBank (http://www.ncbi.nlm.nih.gov) databases (Table 2).
KYPHOSUS VAIGIENSIS RECORDS FROM THE MEDITERRANEAN SEA Zootaxa 3963 (1) © 2015 Magnolia Press · 47
