Low Copy Number DNA Template Can Render Polymerase Chain Reaction Error Prone in a Sequence-Dependent Manner
作者： Mansour Akbari , Marianne Doré Hansen , Jostein Halgunset , Frank Skorpen , Hans E. Krokan
DOI: 10.1016/S1525-1578(10)60006-2
关键词: Low copy number 、 Mutation 、 Inverse polymerase chain reaction 、 DNA 、 Genetics 、 Multiple displacement amplification 、 Uracil-DNA glycosylase 、 Molecular biology 、 DNA clamp 、 Biology 、 Real-time polymerase chain reaction
摘要: Paraffin-embedded tissue is an important source of material for molecular pathology and genetic investigations. We used DNA isolated from microdissected formalin-fixed, paraffin-embedded gastric tumors mutation analysis a region the human gene uracil-DNA glycosylase (UNG), encoding UNG catalytic domain, detected apparent base substitutions which, after further investigation, proved to be polymerase chain reaction (PCR) artifacts. demonstrate that low template input in PCR can generate false mutations, mainly guanine adenine transitions, sequence-dependent manner. One such identical previously reported glioma. This phenomenon was not caused by microheterogeneity sample because same artifact seen amplification homogenous, diluted plasmid. did observe genuine mutations 16 samples. Our results caution should taken when interpreting data PCR-based somatic using amounts DNA, methods enrich putative subpopulations mutant molecules could, essence, PCR-generated
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