Development of D-Loop mitochondrial markers for amplification of prey DNA from wolf scat

Abstract

Analysis of wolves dietary is a currently important theme because of the discussion about wolves preying on livestock as sheep or goats. We developed molecular markers to especially amplify the DNA of the prey out of wolf scat. For this purpose, we used the mitochondrial D-Loop using public available sequences for wolf and seven potential prey species (even-toed ungulates). We developed special primers amplifying either the wolves DNA or the prey DNA. In a fragment of 223-225 basepairs (bp) length we identified 21 SNPs, two 1-bp indels and one 3-bp indel, and three microsatellites to separate seven prey species from each other. Validation of the markers was performed by sequencing the PCR products of 12 fresh prey tissues and 20 wolf scat samples using the different primer pairs.

Overcoming taxonomic challenges in DNA barcoding for improvement of identification and preservation of clariid catfish species

“…Over 500 sequences in classes 2 and 3 were labeled as Clarias sp., Angolian walking catfish ( C. angolensis ), bighead catfish ( C. macrocephalus ), North African catfish ( C. gariepinus ), smoothhead catfish ( C. liocephalus ), Valenciennes clariid ( C. dussumieri ), walking catfish ( C. batrachus ), whitespotted clarias ( C. fuscus ), C. buthupogon , C. camerunensis , C. pachynema, C. gabonensis , and C. magur for COI and Clarias sp., bighead catfish ( C. macrocephalus ), blackskin catfish ( C. meladerma ), blunttooth catfish ( C. ngamensis ), North African catfish ( C. gariepinus ), slender walking catfish ( C. nieuhofii ), walking catfish ( C. batrachus ), whitespotted clarias ( C. fuscus ), C. anguillaris , C. gabonensis , C. kapuasensis , forest walking catfish ( C. leiacanthus ), C. maurus , C. pseudonieuhofii , and C. punctatus for Cytb , suggesting that species identifications in clariid catfish using these markers remain uncertain. However, most of species whose D-loop sequences were examined were categorized into classes 2 and 3, indicating that D-loop sequences are not applicable for identification of clariid catfish species, which is consistent with the findings for D-loop sequences in other vertebrates [ 49 ]. The success rate of DNA barcode identification in clariid catfish is relatively low (91%) compared to that in teleosts, due to the difficulty in detecting errors and confirming taxonomic accuracy or contamination [ 3 , 50 ].…”

“…punctatus for Cytb, suggesting that species identifications in clariid catfish using these markers remain uncertain. However, most of species whose D-loop sequences were examined were categorized into classes 2 and 3, indicating that D-loop sequences are not applicable for identification of clariid catfish species, which is consistent with the findings for D-loop sequences in other vertebrates [49]. The success rate of DNA barcode identification in clariid catfish is relatively low (91%) compared to that in teleosts, due to the difficulty in detecting errors and confirming taxonomic accuracy or contamination [3,50].…”