Diagonsis establishment of fluorescen quantitative PCR assay for pseudorabies wild-type virus and vaccine virus / 生物工程学报
Chinese Journal of Biotechnology
; (12): 1149-1154, 2008.
Article
in Zh
| WPRIM
| ID: wpr-275411
Responsible library:
WPRO
ABSTRACT
We designed two pairs of primers and their corresponding TaqMan probes according to gH, gE gene of PRV. By optimizing the probe's concentration, Mg2+ concentration, primers concentration and sample DNA extraction, real-time fluorescent quantitative PCR (FQ-PCR) which can quickly identity field virus and vaccine virus of PRV was established. According to our results, the dynamic range of the FQ-PCR assay is between 10 x 10(1) copies/microL and 10 x l0(8) copies/microL, and the detection limit of FQ-PCR is 1.0 x 10(1) copies/microL, which is 100 fold higher than that of conventional PCR. We detected 60 doubtful tissue samples using the FQ-PCR assay, serum neutralization and conventional PCR. In conclusion, the FQ-PCR method is rapid, sensitive, specific and accurate, and can be used to detect field strains of PRV rapidly. The closed-tube format of the assay minimized the risk of contamination of subsequent reaction and the assay can be performed in 2 h or less. Development of real-time quantitative PCR provides the basis for the early and rapid detection and analyzing quantitatively the infectious degree of PRV.
Full text:
1
Index:
WPRIM
Main subject:
Pseudorabies
/
Swine
/
Virology
/
Polymerase Chain Reaction
/
Herpesvirus 1, Suid
/
Pseudorabies Vaccines
/
Diagnosis
/
Allergy and Immunology
/
Fluorescent Dyes
/
Genetics
Type of study:
Diagnostic_studies
Limits:
Animals
Language:
Zh
Journal:
Chinese Journal of Biotechnology
Year:
2008
Type:
Article