PMID- 10407470
OWN - NLM
STAT- MEDLINE
DCOM- 19990902
LR  - 20031114
IS  - 0002-9645 (Print)
IS  - 0002-9645 (Linking)
VI  - 60
IP  - 7
DP  - 1999 Jul
TI  - Restriction fragment length polymorphism analysis of strains of porcine 
      reproductive and respiratory syndrome virus by use of a nested-set reverse 
      transcriptase-polymerase chain reaction.
PG  - 802-6
AB  - OBJECTIVE: To increase the timeliness and sensitivity of a procedure that uses 
      viral nucleic acid amplification followed by restriction fragment length 
      polymorphism (RFLP) analysis for identifying strains of porcine reproductive and 
      respiratory syndrome virus (PRRSV). SAMPLE POPULATION: 24 strains of PRRSV. 
      PROCEDURE: A nested-set reverse transcriptase-polymerase chain reaction (RT-PCR) 
      was developed and compared with a nonnested-set RT-PCR for sensitivity in 
      amplifying known quantities of infective PRRSV. Once reaction conditions were 
      optimized, the nested-set RT-PCR was tested for effectiveness with 24 strains of 
      PRRSV isolated from swine. RESULTS: The nested-set RT-PCR was 100- to 1,000-fold 
      more sensitive than the nonnested-set RT-PCR, detecting as little as 1 infective 
      unit of PRRSV/ml of sample. It also was generally as sensitive as the combination 
      of steps, namely virus isolation or propagation and nonnested-set RT-PCR, 
      currently used routinely for amplifying PRRSV prior to RFLP analysis, and it was 
      effective for amplifying all of the 24 strains of PRRSV tested. Using this 
      RT-PCR, all tests were completed within 1.5 days (including RFLP analysis), 
      compared with the > 7 days often required for the currently used method involving 
      virus isolation and propagation. CONCLUSIONS: The nested-set RT-PCR was generally 
      as sensitive as the combination of methods now used for PRRSV amplification prior 
      to RFLP analysis, and it can markedly reduce the time required for testing. 
      CLINICAL RELEVANCE: Presumptive identification of PRRSV strains can be provided 
      in a more timely manner by use of a nested-set RT-PCR.
FAU - Umthun, A R
AU  - Umthun AR
AD  - Virology Swine Research Unit, National Animal Disease Center, USDA, Ames, IA 
      50010, USA.
FAU - Mengeling, W L
AU  - Mengeling WL
LA  - eng
PT  - Journal Article
PL  - United States
TA  - Am J Vet Res
JT  - American journal of veterinary research
JID - 0375011
RN  - 0 (DNA Primers)
RN  - 0 (DNA, Viral)
RN  - 0 (RNA, Viral)
SB  - IM
MH  - Animals
MH  - Bronchoalveolar Lavage Fluid/virology
MH  - Cells, Cultured
MH  - DNA Primers/chemistry
MH  - DNA, Viral/chemistry
MH  - Electrophoresis, Agar Gel/veterinary
MH  - Female
MH  - Open Reading Frames/genetics
MH  - Polymorphism, Restriction Fragment Length
MH  - Porcine Reproductive and Respiratory Syndrome/classification/*diagnosis
MH  - Porcine respiratory and reproductive syndrome 
      virus/chemistry/classification/*genetics
MH  - RNA, Viral/blood
MH  - Reproduction/physiology
MH  - Respiratory Tract Diseases/diagnosis/veterinary
MH  - Reverse Transcriptase Polymerase Chain Reaction/veterinary
MH  - Sensitivity and Specificity
MH  - Specific Pathogen-Free Organisms
EDAT- 1999/07/17 00:00
MHDA- 1999/07/17 00:01
CRDT- 1999/07/17 00:00
PHST- 1999/07/17 00:00 [pubmed]
PHST- 1999/07/17 00:01 [medline]
PHST- 1999/07/17 00:00 [entrez]
PST - ppublish
SO  - Am J Vet Res. 1999 Jul;60(7):802-6.