[Forensic applications of the sequencing of mitochondrial DNA cytochrome oxidase subunit I and II gene for idenfication of Sarcosaphagous flies (diptera)].
OBJECTIVE
This study was based on a 278 and 635 bp region of the gene for cytochrome oxidase subunit I and I (CO I and CO II ) encoding region of mtDNA; the aim was to solve the problems in identifying Sarcosaphagous flies, particularly in the flies' larvae and eggs which could not be identified only by use of their morphologyical features.


METHODS
Samples of sarcosaphagous flies and larvae were collected from those on the corps of rabbits on the grassland in the Huhhot district and of a pig on the sandy ground in the Dunhuang district. The mtDNA of flies and their larvae and eggs was extracted using the Chelex technique. Polymerase chain reactions were conducted on a Perkin-Elmer 9600 thermal cycler, followed by vertical non-denaturing polyacrylamide gelectrophoresis. PCR products were purified using the Nucleic Acid Purification Kit. Sequences of both strands were obtained by direct sequencing of the double-stranded PCR product using one of the PCR primers and the ABI PRISM Big Dye Terminator Cycle Sequencing Kit. Sequence reactions were electrophoresed on ABI Model 377 DNA Sequencers. A neighbour-joining tree using the Tamura and Nei model of nucleotide substitution was constructed using the MEGA2. 1 package.


RESULTS
A 278 and 635 bp region of the gene for CO I and CO II encoding region of mtDNA of Sarcosaphagous flies and their larvae and eggs was noted to show the percentages for the sequence divergence within species (less than 1%) and the sequence divergence between species (above 3%). For species that diverged from all others by a relatively large percentage and had small within-species variation, the least percentages of sequence divergence were given which distinguished any individual within that species from any other species.


CONCLUSIONS
A 278 and 635 bp region of the gene for CO I and CO II encoding region of mtDNA of Sarcosaphagous flies and their larvae and eggs has been effectively used for the molecular phylogeny and the identification of their species group. CO I and CO II, or CO II, is better than CO I.
Cai
Ji-feng
JF
Department of Forensic Pathology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
Liao
Ga
G
Liu
Min
M
Ying
Bin-wu
BW
Zhang
Lin
L
Tao
Tao
T
Deng
Zhen-hua
ZH
Pan
Hong-fu
HF
Deng
Jian-qiang
JQ
Liao
Zhi-gang
ZG
D000818:Animals; D004272:DNA, Mitochondrial / Q000235:genetics*; D004175:Diptera / Q000145:classification / Q000235:genetics*; D003576:Electron Transport Complex IV / Q000235:genetics*; D005554:Forensic Medicine; D006801:Humans; D007814:Larva / Q000235:genetics; D009198:Myiasis / Q000235:genetics*; D010802:Phylogeny; D017422:Sequence Analysis, DNA; D013045:Species Specificity
D004272:DNA, Mitochondrial; D003576:Electron Transport Complex IV