Japanese Journal of Veterinary Research;Volume 66 Number 4


Multi-locus exon-primed intron crossing (EPIC) primer design for regional birds and algorithm design for a combination of introns

Kumakura, Keiko;Osanai, Yuta;Osa, Yuichi;Fujii, Kei;Asakawa, Mitsuhiko;Ushiyama, Kii;Onuma, Manabu;Igarashi, Hirotaka;Endoh, Daiji

Permalink : http://hdl.handle.net/2115/72019
JaLCDOI : 10.14943/jjvr.66.4.261
KEYWORDS : EPIC;polymorphic introns;species-identification;Hokkaido;Max-covering computer program


There are a variety of purposes for identifying the different species of wild animals and birds. It needs a method that can save cost and time for the investigation of many birds. One of which is DNA barcoding that has been used for species identification from sampled muscle, feather or feces. In this study, we developed a novel nuclear exon-primed intron-crossing (EPIC) markers to distinct local birds in Hokkaido area. In birds, the size of introns is smaller than that of mammals. Many introns are within the size range that is suitable for EPIC. To design the EPIC primers, we collated the genome sequences of chicken, turkey, zebra finch and flycatcher on which genes and exons were annotated. We tested the primers for 16 introns which could amplify the introns in 12 different species birds. On the analysis of amplified introns, the size of polymorphisms has been shown in some species of birds. While an intron in TBC1D22B gene showed an interspecies difference in the sizes of amplified introns, some of the size difference was too small for reproducible observation. Since a species could not be identified with one gene, attempts were made to identify species by combining multiple genes. Resultantly, the amplified products of 3 introns have distinguished 10 species of birds. It has been noted that the EPIC method is capable of distinguishing 10 species of local birds in Hokkaido and is therefore suggested to be utilized as a convenient means for multiple species identification.