Producing STR (Short Tandem Repeat) Locus Patterns From Bloodstains and Other Forensic Samples Using an Infrared Fluorescent Automated DNA Sequencer

• NCJ Number: 164335
• Journal: Journal of Forensic Sciences Volume: 41 Issue: 3 Dated: (May 1996) Pages: 418-424
• Author(s): R Roy; D L Steffens; B Gartside; G Y Jang; J A Brumbaugh
• Date Published: 1996
• Length: 7 pages
• Publication Format: Article
• Publication Type: Test/Measurement
• Language: English
• Country: United States of America

Annotation

This research was undertaken to determine if an automated on-line detection system that uses infrared laser irradiation can be used to detect STR (short tandem repeat) alleles from bloodstains with minimal DNA extraction.

Abstract

STR analysis is increasingly being used in forensic case analysis because of the large number of STR loci in the human genome and their highly polymorphic nature. An automated DNA sequencer that uses high-sensitivity infrared fluorescence technology was used to detect STR allele patterns from simulated forensic samples. The amplification strategy used a 19-base-pair extension on the five-foot end of one of the PCR primers. This sequence is identical to the sequence of a universal M13 Forward sequencing primer that is included in the amplification reaction. Allelic bands were detected by incorporation of the M13 primer- fluorescent dye conjugate into PCR products, thus eliminating the need for direct conjugation of fluorescent dye to individual STR primers. By using an infrared-based automated DNA sequencer and Tth DNA polymerase, polymorphic STR alleles were detected on-line rapidly and efficiently from bloodstains by using only a high- temperature incubation to extract DNA from blood cells. Five STR loci were also amplified by using Chelex extracted DNA from simulated forensic samples. Multiplexing of three primer pairs in a single PCR mixture for amplification was accomplished by using Taq polymerase. This system combines infrared fluorescence chemistry and laser technology, thus eliminating the need for radioactivity and the gel handling required with silver staining and fluor detection systems. Real-time detection permits immediate visualization of the data, and STR alleles are displayed as familiar autoradiogramlike images that can be analyzed by computer. By loading a 64-lane gel twice and multiplexing with three primer pairs, forensic scientists can type at least three loci from 120 samples in one day. 6 figures and 23 references