Quantitative polymerase chain reaction assay for largemouth bass virus.
The use of quantitative polymerase chain reaction (QPCR) to test for largemouth bass virus (LMBV) was evaluated during a challenge experiment in which largemouth bass Micropterus salmoides were immersed in the type strain of LMBV. The real-time PCR and cell culture methods were both used to measure LMBV present in the inoculum. Additional samples tested by QPCR included gill, gonad, kidney, liver, mucus, spleen, and swim bladder. A plasmid clone containing a 248-base pair (bp) fragment of the major capsid protein gene (MCP*) was serially diluted and used as a standard to quantify the number of LMBV DNA copies present in the samples tested. A 62-bp fragment of DNA located in MCP* was amplified in the real-time PCR assay. This work has demonstrated the value of the QPCR assay in LMBV surveys.
Getchell
R G
RG
Aquatic Animal Health Program, Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, New York 14853, USA. rgg4@cornell.edu
Groocock
G H
GH
Schumacher
V L
VL
Grimmett
S G
SG
Wooster
G A
GA
Bowser
P R
PR
D000818:Animals; D001492:Bass / Q000821:virology*; D036022:Capsid Proteins / Q000235:genetics; D002460:Cell Line; D017931:DNA Primers / Q000737:chemistry; D004266:DNA Virus Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D005393:Fish Diseases / Q000175:diagnosis* / Q000821:virology; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D017995:Ranavirus / Q000235:genetics / Q000302:isolation & purification*; D012016:Reference Values; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D036022:Capsid Proteins; D017931:DNA Primers