Abstract
Polymerase chain reaction (PCR) is recognized as a rapid, sensitive, and specific molecular diagnostic tool for the analysis of nucleic acids. However, the sensitivity and kinetics of diagnostic PCR may be dramatically reduced when applied directly to biological samples, such as blood and feces, owing to PCR-inhibitory components. As a result, pre-PCR processing procedures have been developed to remove or reduce the effects of PCR inhibitors. Pre-PCR processing comprises all steps prior to the detection of PCR products, that is, sampling, sample preparation, and deoxyribonucleic acid (DNA) amplification. The aim of pre-PCR processing is to convert a complex biological sample with its target nucleic acids/cells into PCR-amplifiable samples by combining sample preparation and amplification conditions. Several different pre-PCR processing strategies are used: (1) optimization of the DNA amplification conditions by the use of alternative DNA polymerases and/or amplification facilitators, (2) optimization of the sample preparation method, (3) optimization of the sampling method, and (4) combinations of the different strategies. This review describes different pre-PCR processing strategies to circumvent PCR inhibition to allow accurate and precise DNA amplification.
| Original language | English |
|---|---|
| Journal | Molecular Biotechnology |
| Volume | 26 |
| Issue number | 2 |
| Pages (from-to) | 133-146 |
| Number of pages | 14 |
| ISSN | 1073-6085 |
| DOIs | |
| Publication status | Published - 2004 |
| Externally published | Yes |
Keywords
- Animals
- Blood Chemical Analysis
- DNA
- Feces
- Humans
- Microchemistry
- Polymerase Chain Reaction
- Specimen Handling
- 9007-49-2 DNA
- antibody
- betaine
- bovine serum albumin
- detergent
- dimethyl sulfoxide
- DNA binding protein
- DNA polymerase
- glycerol
- hemoglobin
- immunoglobulin G
- lactoferrin
- macrogol 400
- organic solvent
- polymer
- polysorbate 20
- aqueous solution
- bacterial growth
- biosynthesis
- blood
- centrifugation
- diagnostic accuracy
- diagnostic procedure
- dilution
- DNA extraction
- DNA synthesis inhibition
- enrichment culture
- feces
- filtration
- gene amplification
- human
- nonhuman
- nucleic acid analysis
- polymerase chain reaction
- review
- sampling
- Bacteria (microorganisms)
- Bovinae
- Amplification facilitators
- PCR facilitators
- PCR inhibitors
- PCR sample
- PCR-amplifiable samples
- PCR-compatible samples
- Pre-PCR processing
- Sample preparation
- Thermostable DNA polymerase
- Primates Mammalia Vertebrata Chordata Animalia (Animals, Chordates, Humans, Mammals, Primates, Vertebrates) - Hominidae [86215] human common
- DNA polymerase 9012-90-2 EC 2.7.7.7
- nucleic acids
- 03502, Genetics - General
- 03508, Genetics - Human
- 10062, Biochemistry studies - Nucleic acids, purines and pyrimidines
- 10802, Enzymes - General and comparative studies: coenzymes
- 14004, Digestive system - Physiology and biochemistry
- 15002, Blood - Blood and lymph studies
- 15004, Blood - Blood cell studies
- Biochemistry and Molecular Biophysics
- blood blood and lymphatics
- feces digestive system
- DNA amplification genetic techniques, laboratory techniques
- polymerase chain reaction genetic techniques, laboratory techniques
- sample preparation laboratory techniques
- sampling laboratory techniques
- Methods and Techniques
- Molecular Genetics