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DNA damage promotes mistyping in the allele specific oligonucleotide probing analysis of forensic samples
Author(s) -
Fattorini Paolo,
Cossutta Federica,
Giulianini Piero,
Edomi Paolo,
Previderè Carlo
Publication year - 2000
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/1522-2683(20000801)21:14<2969::aid-elps2969>3.0.co;2-7
Subject(s) - polymerase chain reaction , oligonucleotide , biology , microbiology and biotechnology , typing , locus (genetics) , allele , str analysis , dna , taq polymerase , genetics , multiple displacement amplification , massive parallel sequencing , dna sequencing , microsatellite , gene , dna extraction , thermus aquaticus
Abstract Five polymerase chain reaction (PCR) products which could not be reliably typed by allele‐specific oligonucleotide (ASO) probing at the human leukocyte antigen (HLA) DQA1 locus were analyzed by polyacrylamide gel electrophoresis and direct sequencing. The first method revealed the preferential amplification of only one of the two alleles in two cases. Direct sequencing of PCR products allowed unambiguous genetic typing but a high number of artifacts was observed. Several of these artifacts occurred in the sequences recognized by the ASOs. This finding provides an explanation for the mistyping in the ASO probing procedure because Taq polymerase errors both created new genetic specificities and eliminated site‐specific polymorphisms. Reversed‐phase HPLC‐MS of the five forensic templates showed a high degree of DNA damage. These data together indicate that the risk of mistyping when using the ASO probing procedure cannot be neglected in the forensic analysis of damaged DNA samples.