DETECTION OF FLAVIVIRUSES BY REVERSE-TRANSCRIPTASE POLYMERASE CHAIN-REACTION

被引:60
作者
ELDADAH, ZA
ASHER, DM
GODEC, MS
POMEROY, KL
GOLDFARB, LG
FEINSTONE, SM
LEVITAN, H
GIBBS, CJ
GAJDUSEK, DC
机构
[1] US FDA, BETHESDA, MD 20014 USA
[2] UNIV MARYLAND, COLLEGE PK, MD 20742 USA
关键词
ST-LOUIS ENCEPHALITIS; JAPANESE ENCEPHALITIS; DENGUE; YELLOW FEVER; REVERSE TRANSCRIPTASE; NESTING PRIMERS; RNA;
D O I
10.1002/jmv.1890330410
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
RNA sequences of five flaviviruses were detected by a modified polymerase chain reaction (PCR) that incorporated a reverse transcriptase and RNase inhibitor. Oligonucleotide primer pairs were synthesized to amplify sequences from St. Louis encephalitis (SLE), Japanese encephalitis (JBE), yellow fever (YF), dengue 2 (DEN-2), and dengue 4 (DEN-4) viruses. The amplified products were visualized as bands of appropriate size on ethidium bromide-stained agarose gels. The identity of these products was confirmed by restriction endonuclease cleavage to generate fragments of predicted lengths. The reverse-transcriptase PCR (RT-PCR) successfully amplified flavivirus sequences from cell cultures, frozen brain tissue, and formalin-fixed, paraffin-embedded brain tissue. The reactions were highly specific, and the method compared favorably to two conventional assays of viral infectivity. RT-PCR followed by PCR with nesting primers (N-PCR) was 1,000-fold more sensitive in detecting virus than classical infectivity titration by intracerebral inoculation of suckling mice and nearly 1,000-fold more sensitive than amplification of virus in cell culture followed by inoculation of mice.
引用
收藏
页码:260 / 267
页数:8
相关论文
共 29 条
[1]   RAPID AND SIMPLE METHOD FOR PURIFICATION OF NUCLEIC-ACIDS [J].
BOOM, R ;
SOL, CJA ;
SALIMANS, MMM ;
JANSEN, CL ;
WERTHEIMVANDILLEN, PME ;
VANDERNOORDAA, J .
JOURNAL OF CLINICAL MICROBIOLOGY, 1990, 28 (03) :495-503
[2]   ISOLATION OF BIOLOGICALLY-ACTIVE RIBONUCLEIC-ACID FROM SOURCES ENRICHED IN RIBONUCLEASE [J].
CHIRGWIN, JM ;
PRZYBYLA, AE ;
MACDONALD, RJ ;
RUTTER, WJ .
BIOCHEMISTRY, 1979, 18 (24) :5294-5299
[3]  
CRISTIANO K, 1991, IN PRESS HEPATOLOGY
[4]  
DAVIS LG, 1986, BASIC METHODS MOL BI, P130
[5]   A COMPREHENSIVE SET OF SEQUENCE-ANALYSIS PROGRAMS FOR THE VAX [J].
DEVEREUX, J ;
HAEBERLI, P ;
SMITHIES, O .
NUCLEIC ACIDS RESEARCH, 1984, 12 (01) :387-395
[6]   AMPLIFICATION OF RHINOVIRUS SPECIFIC NUCLEIC-ACIDS FROM CLINICAL-SAMPLES USING THE POLYMERASE CHAIN-REACTION [J].
GAMA, RE ;
HORSNELL, PR ;
HUGHES, PJ ;
NORTH, C ;
BRUCE, CB ;
ALNAKIB, W ;
STANWAY, G .
JOURNAL OF MEDICAL VIROLOGY, 1989, 28 (02) :73-77
[7]   DETECTION OF MEASLES-VIRUS GENOMIC SEQUENCES IN SSPE BRAIN-TISSUE BY THE POLYMERASE CHAIN-REACTION [J].
GODEC, MS ;
ASHER, DM ;
SWOVELAND, PT ;
ELDADAH, ZA ;
FEINSTONE, SM ;
GOLDFARB, LG ;
GIBBS, CJ ;
GAJDUSEK, DC .
JOURNAL OF MEDICAL VIROLOGY, 1990, 30 (04) :237-244
[8]   GENERATION OF SINGLE-STRANDED-DNA BY THE POLYMERASE CHAIN-REACTION AND ITS APPLICATION TO DIRECT SEQUENCING OF THE HLA-DQA LOCUS [J].
GYLLENSTEN, UB ;
ERLICH, HA .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1988, 85 (20) :7652-7656
[9]   NUCLEOTIDE-SEQUENCE OF DENGUE-2 RNA AND COMPARISON OF THE ENCODED PROTEINS WITH THOSE OF OTHER FLAVIVIRUSES [J].
HAHN, YS ;
GALLER, R ;
HUNKAPILLER, T ;
DALRYMPLE, JM ;
STRAUSS, JH ;
STRAUSS, EG .
VIROLOGY, 1988, 162 (01) :167-180
[10]   HUMAN INFECTION ACQUIRED IN LABORATORY [J].
HAMMON, WM .
JOURNAL OF THE AMERICAN MEDICAL ASSOCIATION, 1968, 203 (09) :647-&