Computational protocol: Two gonad-infecting species of Philometra (Nematoda: Philometridae) from groupers (Serranidae) off Tunisia, with a key to Philometra species infecting serranid gonads

Similar protocols

Protocol publication

[…] Fish were purchased at the fish market in Tunis and Sfax, Tunisia; these were previously caught by fishermen in the nearby coastal waters of the Mediterranean Sea. Fish DNA was extracted from tissue samples using the NucleoSpin 96 tissue kit (Macherey-Nagel, Düren, Germany) following the manufacturer’s instructions. Sequences were obtained by amplification and sequencing of a region of the cytochrome oxidase subunit I (COI) mitochondrial gene using the primers FishF1 (5′-TCAACYAATCAYAAAATYGGCAC-3′) and FishR1 (5′-TGATTYTTYGGYCACCCRGAAGT-3′) []. Standard PCRs were carried out in 20 μL total volume, containing about 30 ng of DNA, 1 × 10× PCR buffer, 2 mM MgCl2, 200 μM mix dNTPs, 150 nM of each primer and 1 unit of Taq polymerase (Qiagen, Hilden, Germany). After an initial denaturation of 3 min at 95 °C, the mitochondrial DNA was amplified through 39 cycles of 15 s at 95 °C, 20 s at 48 °C and 40 s at 72 °C, with a terminal elongation for 5 min at 72 °C. PCR products were purified and sequenced in both directions on 3730xl DNA Analyser 96-capillary sequencer (Applied Biosystems, Waltham, MA, USA). Sequences were edited using CodonCode Aligner software (CodonCode Corporation, Dedham, MA, USA), compared with the GenBank database content using BLAST, and deposited in GenBank under Accession Numbers KU739518–KU739521. Species identification was confirmed using the BOLD identification engine []. Since BOLD does not include all sequences available in GenBank but includes others, comments are added for similarities with other sequences. The fish nomenclature adopted follows FishBase []. […]

Pipeline specifications

Software tools FISHR, CodonCode Aligner
Databases FishBase
Application Population genetic analysis
Organisms Caenorhabditis elegans, Nitrosomonas sp.