Chemiluminescent detection of RFLP patterns in forensic DNA analysis.
DNA testing by restriction fragment length polymorphism (RFLP) analysis is an extremely important technique used in forensic science laboratories. While RFLP testing is a highly informative method, it traditionally has had several disadvantages. It is time consuming and involves work with radioactive phosphorous. A detection method that is faster and safer than isotopic detection is presented. Various membranes, fixation methods and transfer procedures were evaluated for DNA retention and sensitivity using alkaline phosphatase conjugated oligonucleotide probes and a chemiluminescent substrate. Blood samples and evidentiary material from forensic casework were analyzed by both chemiluminescent and isotopic detection. Results of each method were compared for pattern appearance, band size, and composite profile frequency. The chemiluminescent system had very good sensitivity, detecting 3-25 ng K562 DNA. Most patterns developed by both methods appeared the same. The variation observed between band sizes and frequency estimates generated by each method was as expected for an inter-gel comparison. The chemiluminescent detection procedure described here is suitable for use in forensic casework.
Johnson
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U.S. Army Criminal Investigation Laboratory, Fort Gillem, GA, USA.
Kotowski
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D000483:Alleles; D001801:Blood Stains; D015342:DNA Probes / Q000032:analysis*; D005554:Forensic Medicine / Q000379:methods*; D006801:Humans; D008163:Luminescent Measurements; D008970:Molecular Weight; D012150:Polymorphism, Restriction Fragment Length; D012680:Sensitivity and Specificity
D015342:DNA Probes