Identifying PCR primers to facilitate molecular phylogenetics in Caddisflies (Trichoptera)

Abstract

The molecular phylogenetics of the Lepidoptera (butterflies and moths) is well studied, but that of Trichoptera (caddisflies), the sister clade of Lepidoptera, is less studied. The PCR primer libraries developed for lepidopteran phylogenetics might work in Trichoptera. DNA from 8 caddisfly species (Asynarchus nigriculus (Banks, 1908), Grammotaulius lorettae Denning, 1941, Hesperophylax occidentalis (Banks, 1908), Limnephilus externus Hagen, 1861, Limnephilus picturatus McLachlan, 1875, Limnephilus secludens Banks, 1914, Limnephilus sublunatus Provancher, 1877 and Agrypnia deflata (Milne, 1931)) was used to screen for amplification. 107 primer pairs for 45 nuclear and 3 mitochondrial genes were tested. Primers for 1 new gene (40S ribosomal protein S2 (RPS2)) and 8 genes previously used in Trichopteran phylogenetics were recovered (16S rRNA, 18S rRNA, carbamoyl-phosphate synthetase (CAD), cytochrome oxidase I (COI), cytochrome oxidase II (COII), elongation factor-1 alpha (EF-1 alpha), isocitrate dehydrogenase (IDH), and RNA polymerase-II (POL-II)). New primer pairs extended the genomic region sampled for many genes. Evolution rates among loci varied by 2 orders of magnitude. Differences among evolution rates and modes of inheritance offer flexible tools for resolving phylogenetic questions and examining genome evolution in the Trichoptera. Screening libraries of PCR primers is a useful approach for identifying PCR primers in related taxa with limited molecular genetic resources.