A combination of techniques proves useful in the development of nuclear markers in the newt genus Triturus

Abstract

To increase the number of markers available for study of phylogeny and phylogeography in the newt genus Triturus, we developed and tested 59 primer pairs using three different techniques. Primers were obtained from published sources, by designing exon-primed intron-crossing primers and from randomly cloned anonymous nuclear DNA fragments. Successful polymerase chain reaction products were cloned and sequenced. Five fragments were successfully amplified and sequenced for six species of Triturus: intron 7 of the β-fibrinogen gene (βfibint7), third intron of the calreticulin gene (CalintC), the 11th intron of the α-subunit of the platelet derived growth factor receptor (PDGFRα) and two anonymous markers (Cri1 and Cri4). The average percentage species divergence across all the markers is low (c. 3%), compared to what has been found in mitochondrial DNA (25-30%).