Tandem duplication of mitochondrial DNA in the black-faced spoonbill, Platalea minor

Mitochondrial (mt) heteroplasmy in the control region (CR) of the black-faced spoonbill was investigated using LA-PCR. To avoid amplification of transpositioned nuclear genome fragment from mtDNA (numt), PCR product of the almost-complete mitochondrial genome was amplified using primers designed to anneal on the COIII gene. Then nested LA-PCR product was amplified between the cyt b and 12S rRNA genes using the almost-complete mitochondrial genome PCR product as a template. Nucleotide sequencing revealed tandem duplication composed of two units. The first contains cyt b-1, tRNA(Thr)-1, tRNA(Pro)-1, ND6-1, tRNA(Glu)-1 and CR1, and the second consists of cyt b-2, tRNA(Thr)-2, tRNA(Pro)-2, ND6-2, tRNA(Glu)-2 and CR2, followed by tRNA(Phe) and 12S rRNA. The duplicated cyt b-2 sequence coincided with 499 bp at the 3' end of cyt b-1. With the exception of the CR, the other genes in the duplicated sequence were identical to the original corresponding gene. Even though both CR1 and CR2 contain functional blocks, such as a poly-C site, a goose hairpin and a TAS structure in Domain I, the 3' end of CR1 was followed by a 112 bp sequence (non-coding region) that was not found in CR2 or in sequence homology analysis of similar genes. Meanwhile, CR2 ended in a complicated repeat sequence. The 5' franking region in the Domain I (Region A) and the 3' franking region in the Domain I (Region B) of the two CRs evolve in quite different manners: Region A was highly variable between CR1 and CR2 in the same individuals, while Region B was almost identical between them, which indicates concerted evolution.