PCR profile was 5 min initial denaturation at 95 ☌ 35 cycles of 30 s denaturation at 95 ☌, 30 s annealing at 50–55 ☌, and 1–2 min extension at 72 ☌ and a final extension at 72 ☌ for 8 min and 4 ☌ hold. PCR was performed in a 30 μL system, which contained 2 × Taq PCR SuperMix (Tiangen, China) 15 μL, 10 μM of each primer (forward and reverse) 1 μL, 1 μL template DNA and 12 μL sterile double-distilled water (ddH2O).
#CLC GENOMICS WORKBENCH 8.5 EXCLUDE HOST USING PRIMERS SOFTWARE#
Sequences obtained were aligned and edited using the software SeqMan (DNAStar, Inc.) to generate complete mitochondrial DNA sequences.Īll primers used in this study were taken from Hu et al. Then the PCR products were purified using a gel extraction kit (Promega) and sequenced with each of the PCR primers on an ABI 377 sequencer. ostralegus was generated by amplification of overlapping polymerase chain reaction (PCR) fragments ( Hu et al. The quality of DNA was assessed through electrophoresis in a 1% agarose gel and staining with Gold View. Total genomic DNA was extracted using standard phenol-chloroform methods ( Sambrook and Russell, 1989). After collection, the tissue was initially preserved in 95% ethanol in the field, and transferred to −80☌ in our laboratory for long-term storage at Nanjing Normal University (specimen voucher number: NJNU-Host07). ost ralegus was collected from a derelict and abandoned mist net in Dongtai, Yancheng City, Jiangsu Province, China (32☄6’23.12” N, 120★7’41.68” E), in July 2017. Our experimental procedures complied with the current laws on animal welfare and research in China, and were specifically approved by Nanjing Normal University’s Animal Care and Use Committee (Permit #8). ostralegus, and provide useful information for understanding the evolutionary and taxonomic research within Charadriiformes. ostralegus, which may provide a valuable resource facilitating further study of population genetics and biogeography of H. In this study, we sequenced the complete mitogenome of H. During the past decade, mitogenome provides a valuable resource for further study of molecular systematics, species identification, population genetics, phylogeny, taxonomy and so on ( Li et al. ostralegus.Īnimal mitogenome typically contains 13 protein-coding genes, 2 ribosomal RNAs (12S rRNA and 16S rRNA), 22 transfer RNAs (tRNAs), and a non-coding control region ( Ruokonen and Kvist, 2002). Few population genetics have been carried out, which might be the lack of basic genetics data (or molecular markers) of H. Based on eight polymorphic microsatellite loci, no significant genetic differentiation was observed between two groups of ‘residents’ and ‘leapfrogs’, which were divided by the observational data on the dispersal behaviour of breeding individuals in the island of Schiermonnikoog, Netherlands ( Van et al. ostralegus pay more attention on habitat ecology, foraging behavior ( Schwemmer et al. Eurasian oystercatcher feeds on mussels, crabs, earth worms and all kinds of invertebrates, rarely on fish. This species has been categorized as near threatened in red list of threatened species. The Eurasian oystercatcher, Haematopus ostralegus (Charadriiformes, Haematopodidae), is a relatively large wader with an extremely large distribution ranging from Europe to Siberia. , 2007 Gibson and Baker, 2012 Friesen, 2015 Hu et al. The resources of mitogenome have rapidly accumulated in recent years thanks to the advanced genomic sequencing, while the mitogenome has not been well studied in Charadriiformes ( Baker et al. This study provides a valuable resource facilitating further study of phylogenetic and evolutionary analysis for wader, and improves our understanding of the evolutionary and taxonomic research within Charadriiformes. Evidence of positive selection was obtained for three (ND2, ND4 and ND6) genes by at least one of the methods. The ratio of Ka/Ks of all PCGs indicated that all these genes evolved under purifying selection. We found a significant negative correlation between CBI and ENC, and a significant positive correlation was found between CBI and G + Cc and G + C3s. In Charadriiformes mitogenomes, AT skews values were positive, while the values of GC skew were negative. ostralegus is a circular molecule of 16,798 bp in length and the overall nucleotide composition of H-strand was A: 31.45%, T: 23.46%, C: 31.30%, G: 13.79%. ostralegus, and illustrating mitogenomes structure and investigating their evolutionary relationship by comparing 31 species in Charadriiformes.
The objective of this study was to determine the complete mitogenome sequence of H. T he Eurasian oystercatcher Haematopus ostralegus is a relatively large migratory wader with an extremely large distribution range.