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Targeting optimal introns for phylogenetic analyses in non‐model taxa: experimental results in Asian pitvipers
Author(s) -
Creer Simon,
Malhotra Anita,
Thorpe Roger S.,
Pook Catharine E.
Publication year - 2005
Publication title -
cladistics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.323
H-Index - 92
eISSN - 1096-0031
pISSN - 0748-3007
DOI - 10.1111/j.1096-0031.2005.00072.x
Subject(s) - biology , phylogenetic tree , intron , genetics , taxon , locus (genetics) , primer (cosmetics) , low copy number , epic , polymerase chain reaction optimization , evolutionary biology , polymerase chain reaction , gene , genome , multiplex polymerase chain reaction , art , chemistry , botany , literature , organic chemistry
Nuclear introns are increasingly used as phylogenetic markers. Here, we present a multidisciplinary approach towards optimal locus selection and amplification using Asian pitvipers as an example of a non‐model taxon, and raise the profile of length variant heterozygotes (LVHs) in intron loci. Taxon‐specific primers were identified using a bioinformatic approach, and also designed from existing exon primed, intron crossing (EPIC) primer amplifications. Eleven further universal EPIC primer pairs were assayed using a range of PCR optimization strategies. Taxon‐specific primers yielded the most consistent amplifications, but assaying a large number of universal EPIC primers yielded another appropriate locus for phylogenetic purposes. Modified Taq DNA polymerases such as JumpStart™ Taq either significantly improved the specificity and yield of EPIC PCR amplifications (of low copy number nuclear targets), or resulted in amplifications that were not significantly worse than those derived from a generic Taq DNA polymerase. Finally, LVHs were detected in all loci that were sequenced suggesting that they are relatively common in introns. This study provides an efficient and cost effective template for the successful identification of intron markers for molecular systematics which is universally applicable to other non‐model taxon groups. © The Willi Hennig Society 2005.