PMID- 9395141
OWN - NLM
STAT- MEDLINE
DCOM- 19980217
LR  - 20190831
IS  - 0166-0934 (Print)
IS  - 0166-0934 (Linking)
VI  - 68
IP  - 1
DP  - 1997 Oct
TI  - Development of a RT-PCR test coupled with a microplate colorimetric assay for the 
      detection of a swine Arterivirus (PRRSV) in boar semen.
PG  - 65-80
AB  - Transmission of porcine reproductive and respiratory syndrome virus (PRRSV) 
      through boar semen has been demonstrated, stressing the need for a reliable semen 
      PRRSV detection test. A diagnostic assay was developed based on amplification of 
      the PRRSV RNA by reverse transcription and polymerase chain reaction (RT-PCR) 
      followed by detection of the amplification products by hybridization and 
      colorimetric assay in microwell plates. A highly reproducible and efficient 
      method of viral RNA isolation from semen samples was set up. A combined RT-PCR 
      procedure was performed, incorporating the use of uracil-N-glycosylase (UNG) in 
      combination with dUTP instead of dTTP to prevent false positive results due to 
      carry-over contamination. An RNA internal control was added during the RNA 
      isolation procedure to detect false negative results. The colorimetric detection 
      in microwell plates of amplification products from either PRRSV or IC RNA gave 
      specific and objective results and was automated. A cut-off value of 1000 RNA 
      copies or 10 TCID50 of PRRSV per ml of semen samples could be detected with this 
      assay. Semen samples collected from experimentally-infected boars were tested 
      with this assay and showed PRRSV excretion early after infection and for an 
      extended period.
FAU - Legeay, O
AU  - Legeay O
AD  - Centre National d'Etudes Veterinaire et Alimentaire, Unite biologie moleculaire, 
      Ploufragan, France.
FAU - Bounaix, S
AU  - Bounaix S
FAU - Denis, M
AU  - Denis M
FAU - Arnauld, C
AU  - Arnauld C
FAU - Hutet, E
AU  - Hutet E
FAU - Cariolet, R
AU  - Cariolet R
FAU - Albina, E
AU  - Albina E
FAU - Jestin, A
AU  - Jestin A
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - Netherlands
TA  - J Virol Methods
JT  - Journal of virological methods
JID - 8005839
RN  - 0 (DNA, Viral)
RN  - 0 (RNA, Viral)
SB  - IM
MH  - Animals
MH  - Base Sequence
MH  - Cloning, Molecular
MH  - Colorimetry/*methods
MH  - DNA, Viral
MH  - Evaluation Studies as Topic
MH  - Genome, Viral
MH  - Male
MH  - Molecular Sequence Data
MH  - Polymerase Chain Reaction/*methods
MH  - Porcine Reproductive and Respiratory Syndrome/immunology/virology
MH  - Porcine respiratory and reproductive syndrome 
      virus/genetics/immunology/*isolation & purification
MH  - RNA, Viral/analysis
MH  - Reproducibility of Results
MH  - Semen/*virology
MH  - Sensitivity and Specificity
MH  - Swine
MH  - Transcription, Genetic
EDAT- 1997/12/12 00:00
MHDA- 1997/12/12 00:01
CRDT- 1997/12/12 00:00
PHST- 1997/12/12 00:00 [pubmed]
PHST- 1997/12/12 00:01 [medline]
PHST- 1997/12/12 00:00 [entrez]
AID - S0166093497001109 [pii]
AID - 10.1016/s0166-0934(97)00110-9 [doi]
PST - ppublish
SO  - J Virol Methods. 1997 Oct;68(1):65-80. doi: 10.1016/s0166-0934(97)00110-9.