Download Assiut university researches Rapid detection of BoHV

Assiut university researches
Rapid detection of BoHV-1 genomic DNA by
loop-mediated isothermal amplification assay
Alaa A. El-Kholya, Khaled Abdelrahmanb, Hatem Solimanc,
Abstract:
Bovine herpes virus-1 (BoHV-1) is a serious viral pathogen of
domestic and wild cattle. Herein, we report development of a
new molecular diagnostic assay for rapid and sensitive
detection of BoHV-1 utilizing the loop-mediated isothermal
amplification (LAMP) technique. BoHV-1-LAMP assay was
optimized to amplify the target DNA by incubation the BstDNA polymerase enzyme with a set of specially constructed
six primers, based on the gE-gene of BoHV-1 virus, at 65 ◦C
for 60 min. BoHV-1-LAMP products were detected by visual
inspection using SYBR Green-I stain and had a ladder-like
appearance by gel electrophoresis analysis. Negative results
obtained with DNA from other tested fish viruses confirmed
the specificity of the assay. The analytical sensitivity of the
BoHV-1-LAMP assay was 1 fg of BoHV-1 DNA (dilution of
106). The developed assay could successfully detect BoVH-1
DNA from clinical samples. Results of this study indicate that
the developed BoHV-1-LAMP is rapid and highly sensitive
assay not only for detection of BoHV-1 in clinical samples, but
also for differentiation between wild-type (gE-positive) and
gE-negative BoHV-1 viruses, which will improve the control
programs of BoHV-1 in Egypt. © 2014 Elsevier B.V. All rights
reserved.
Key words:
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Published in:
Journal of Virological Methods,204,81–85