RAPD analysis and sequence alignment of genomic DNA of hair follicle mites Demodex folliculorum and D. brevis (Acari:Demodicidae)

[Objective] Analysis of genomic DNA polymorphism and the related sequence of Demodex folliculorum and D.brevis.[Methods] The genomic DNA of the human Demodex was extracted by using improved DNA extraction method of mini-insects.Random amplified polymorphic DNA(RAPD)was applied to analyze the polymorphism.The related bands were connected with pMD18-T vector,and cloned,sequenced,and identified and analyzed after enzyme digestion.[Results] There were 15 bands obtained in D.folliculorum and 12 in D.brevis.Some bands were shared by the two mites while others were species-specific.The genetic distance between the two Demodex species was 0.5556.After recombination,the sequence of the specific band(about 800 bp)of D.folliculorum was found to be 855 bp(GenBank accession no.FI277970)in size.The 855 bp fragment was confirmed to be characteristic for D.folliculorum according to the PCR result with a specific primer and the result of enzyme digestion analysis.This sequence had 46% similarity to AraC-type DNA-binding domain-containing protein.The shared fragments(about 300 bp)by the two mites were both 341 bp in length(GenBank accession no.FI520176 and FI520175,respectively),and with two different bases at the 84th and 165th sites,which were A/G and C/T permutation,respectively.Their homology was 99.4%,but there was no ORF and high similar sequence.[Conclusions] The 855 bp fragment is specific to D.folliculorum.The 341 bp fragment is shared by D.folliculorum and D.brevis with 99.4% homology.RAPD can be used in genomic DNA polymorphism analysis and species identitfication of D.folliculorum and D.brevis.