Korean J Vet Res > Volume 55(2); 2015 > Article
Korean Journal of Veterinary Research 2015;55(2):105-110.
DOI: https://doi.org/10.14405/kjvr.2015.55.2.105    Published online June 24, 2015.
Identification of Brucella melitensis isolates originating from Mongolia and diagnostic real-time PCR evaluation using a specific SNP
Sung-Il Kang, Ji-Yeon Kim, Suk Mi Kim, Jin Ju Lee, So-Ra Sung, Yeon-Hee Kim, Suk Chan Jung, Moon Her
OIE Reference Laboratory for Brucellosis, Division of Bacterial Disease, Animal and Plant Quarantine Agency
몽골 유래 Brucella melitensis 동정 및 특이 SNP를 이용한 real-time PCR법에 의한 진단 평가
강성일, 김지연, 김숙미, 이진주, 성소라, 김연희, 정석찬, 허문
농림축산검역본부 세균질병과 OIE 브루셀라 표준연구실
Abstract
A real-time PCR assay using hybridization probe (HybProbe) has been developed to detect Brucella (B.) melitensis strains. The primer and HybProbe sets were designed based on the gap gene of chromosome I with a specific single nucleotide polymorphism of B. melitensis. Specificity of the assay was confirmed by comparison to reference Brucella species and other related strains. In the melting curve analysis, B. melitensis generated a peak at $67^{circ}C$ unlike those for other Brucella species observed at $61^{circ}C$. Sensitivity of the assay for B. melitensis ranged from 20 ng to 200 fg of genomic DNA. The ability to identify 94 Mongolian B. melitensis isolates using the real-time PCR assay was identical to that of classical biotyping methods and differential multiplex PCR. These data showed that this new molecular technique is a simple and quick method for detecting B. melitensis, which will be important for the control and prevention of brucellosis.
Key Words: Brucella melitensis, HybProbe, melting curve, real-time PCR
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