Large-Scale Survey of Mitochondrial D-Loop of the Red-Crowned Crane <i>Grus japonensis</i> in Hokkaido, Japan by Convenient Genotyping Method

Abstract

ABSTRACT. The Red-crowned Crane, Grus japonensis, is an endangered species of crane that has two separate breeding populations, one in the Amur River basin (continental population) and the other in eastern or northern Hokkaido, Japan (island population). So far, only two haplotypes (Gj1 and Gj2) have been identified in the mitochondrial D-loop in island population, whereas seven haplotypes have been found in continental population (Gj3-Gj9). We developed a rapid and inexpensive method of extensive genotyping of D-loop haplotypes in Red-crowned Cranes, based on amplification refractory mutation system (ARMS) PCR assay. Two hundred and three cranes in eastern Hokkaido were studied with this method and supplemental DNA sequencing. Only two haplotypes, Gj1 and Gj2, were confirmed in eastern Hokkaido with Gj2 as a major haplotype. Additionally, only Gj2 was identified in twelve feathers from both sexes found in northern Hokkaido. These results suggest scarce genetic diversity in island population of Red-crowned Cranes in Hokkaido, Japan.

Origin of Three Red-Crowned Cranes <i>Grus japonensis</i> Found in Northeast Honshu and West Hokkaido, Japan, from 2008 to 2012

“…Only two haplotypes (Gj1 and Gj2) were identified in 15 wild cranes from the island population, while the other seven haplotypes (Gj3-Gj9) were detected in only 14 captive cranes in some zoos and aquariums in Japan, birds that originated from the continental population [3]. The finding of extremely low genetic diversity in the island population was confirmed by large-scale study with 203 wild cranes in our previous study [9] as well as a pilot study with microsatellites [2].…”

“…For comparison, body feathers were also obtained from 52 adult Red-crowned Cranes kept in Okayama Prefectural Nature Conservation Center in Wake, Okayama Prefecture in 2009. We used the same samples as those in our previous study [9]. Forty five of these captive cranes originated from birds imported from Eurasian Continent (Gj3, 35: Gj4, 1: Gj5, 1), and 8 cranes were from Hokkaido (Gj2) [9].…”

“…We used the same samples as those in our previous study [9]. Forty five of these captive cranes originated from birds imported from Eurasian Continent (Gj3, 35: Gj4, 1: Gj5, 1), and 8 cranes were from Hokkaido (Gj2) [9].…”

“…Mitochondrial DNA was extracted from several pieces of feather follicles as previously described [9]. Extracted DNA was used as a template for polymerase chain reaction (PCR) to produce the whole length of the D-loop with the forward primer 5′-ACCCCATAATACGGTGAAGG-3′ and the reverse primer 5′-ACGGTAAGGTTAGGACTAAG-3′.…”

“…Complete D-loop was sequenced with the dideoxy method *CorrespondenCe to: Teraoka, H., School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu 069-8501, Japan. e-mail: hteraoka@rakuno.ac.jp ©2013 The Japanese Society of Veterinary Science as described [9]. The primers used for sequencing were SE-QDLF1 (5′-GGCTTTTCTCCAAAACTCGC-3′: forward), SEQDLF2 (5′-TCACGTGAAATCAGCAACCG-3′: forward) and SEQDLR (5′-ATCTTCAGTGCCATGCTTTG-3′: reverse).…”