Biomaterial Database

Quantitative polymerase chain reaction for hepatitis B virus DNA. 1994

J Wu, and D E Sullivan, and M A Gerber

Department of Pharmacology, Tulane University School of Medicine, New Orleans, LA.

To monitor and compare the effect of drugs on hepatitis B virus (HBV) replication in hepatitis patients, an accurate quantitative method with high sensitivity is needed. Polymerase chain reaction (PCR) is the most sensitive method for the detection of HBV DNA, but because of sample to sample variability in amplification efficiency, the quantification of target nucleic acids by conventional PCR is inaccurate. Therefore, we developed a competitive PCR method with an internal standard which uses the same primers as the target HBV DNA. The standard was generated from the HBV S gene by PCR site-directed mutagenesis and differed from the native S gene by a single base which introduced an internal restriction enzyme site. For practical application, this method was used to quantitate the effect of the carbocyclic analogue 2'-deoxyguanosine (2'-CDG) on HBV DNA replication in 2.2.15 cells, an HBV transfected HepG2 cell line. A decrease of HBV DNA level in the attomole range was demonstrated by the assay in the media of 2.2.15 cells treated with 2'-CDG. The results were verified by conventional HBV DNA slot blot analysis followed by densitometric scanning. Competitive PCR proved to be a sensitive, accurate and reliable method for HBV quantitation.

UI	MeSH Term	Description	Entries
D008969	Molecular Sequence Data	Descriptions of specific amino acid, carbohydrate, or nucleotide sequences which have appeared in the published literature and/or are deposited in and maintained by databanks such as GENBANK, European Molecular Biology Laboratory (EMBL), National Biomedical Research Foundation (NBRF), or other sequence repositories.	Sequence Data, Molecular,Molecular Sequencing Data,Data, Molecular Sequence,Data, Molecular Sequencing,Sequencing Data, Molecular
D012015	Reference Standards	A basis of value established for the measure of quantity, weight, extent or quality, e.g. weight standards, standard solutions, methods, techniques, and procedures used in diagnosis and therapy.	Standard Preparations,Standards, Reference,Preparations, Standard,Standardization,Standards,Preparation, Standard,Reference Standard,Standard Preparation,Standard, Reference
D002460	Cell Line	Established cell cultures that have the potential to propagate indefinitely.	Cell Lines,Line, Cell,Lines, Cell
D003849	Deoxyguanosine	A nucleoside consisting of the base guanine and the sugar deoxyribose.
D004279	DNA, Viral	Deoxyribonucleic acid that makes up the genetic material of viruses.	Viral DNA
D006515	Hepatitis B virus	The type species of the genus ORTHOHEPADNAVIRUS which causes human HEPATITIS B and is also apparently a causal agent in human HEPATOCELLULAR CARCINOMA. The Dane particle is an intact hepatitis virion, named after its discoverer. Non-infectious spherical and tubular particles are also seen in the serum.	Dane Particle,Hepatitis Virus, Homologous Serum,B virus, Hepatitis,Hepatitis B viruses,Particle, Dane,viruses, Hepatitis B
D006801	Humans	Members of the species Homo sapiens.	Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000998	Antiviral Agents	Agents used in the prophylaxis or therapy of VIRUS DISEASES. Some of the ways they may act include preventing viral replication by inhibiting viral DNA polymerase; binding to specific cell-surface receptors and inhibiting viral penetration or uncoating; inhibiting viral protein synthesis; or blocking late stages of virus assembly.	Antiviral,Antiviral Agent,Antiviral Drug,Antivirals,Antiviral Drugs,Agent, Antiviral,Agents, Antiviral,Drug, Antiviral,Drugs, Antiviral
D001483	Base Sequence	The sequence of PURINES and PYRIMIDINES in nucleic acids and polynucleotides. It is also called nucleotide sequence.	DNA Sequence,Nucleotide Sequence,RNA Sequence,DNA Sequences,Base Sequences,Nucleotide Sequences,RNA Sequences,Sequence, Base,Sequence, DNA,Sequence, Nucleotide,Sequence, RNA,Sequences, Base,Sequences, DNA,Sequences, Nucleotide,Sequences, RNA
D012680	Sensitivity and Specificity	Binary classification measures to assess test results. Sensitivity or recall rate is the proportion of true positives. Specificity is the probability of correctly determining the absence of a condition. (From Last, Dictionary of Epidemiology, 2d ed)	Specificity,Sensitivity,Specificity and Sensitivity