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A study of DNA identification using nonradioactive automated minisatellite variant repeat (MVR) digital typing
Abstract
The Minisatellite Variant Repeat (MVR) typing technique developed by Dr. Alec Jeffreys (Nature 354:204-209, 1991) has had a significant impact on family studies and forensics because its digital approach can overcome many of the limitations of current DNA typing systems. The purpose of this study was the automation of this technique by using the LI-COR Model 4000 DNA sequencer. Two loci, MS32 (chromosome 1) and MS31 (chromosome 7) were analyzed. About 100ng of human genomic DNA was amplified using a protocol similar to the Jeffreys' MVR typing technique, but with the following changes: (1) A modified 32D primer for MS32 or a 20D for MS31 was used for infrared tagging of the defined repeat segments. (2) A two stage polymerase chain reaction (PCR) was used to increase the specificity and yield of PCR product. (3) A denaturing polyacrylamide gel of only 8cm separation length instead of an agarose gel was used for separation. (4) Data were collected on-line in real time in a continuous fashion with the infrared detector of the LI-COR Model 4000 DNA sequencer. A total of 108 individuals were tested and their identifying diploid codes were determined. An average of 45 repeat units for MS32 and 55 repeat units for MS31 were resolved for each individual. Twenty-eight nuclear families (MS32) and 12 nuclear families (MS31) were evaluated for identification of parentage. Digital patterns from mother and father correctly identified each child from the total pool with MS32 and MS31. The average number of exclusions were 9.6 with a range of 2-19 at MS32 and 15.7 with a range of 5-22 at MS31. Additionally, samples obtained from Operation Desert Storm casualties, two families with identical twins, a family with incidence of Huntington's disease, a family with 5 children, and 8 different primates were tested with either or both loci. Because this method eliminates radioactivity, the time from raw sample to MVR code is approximately 9 hours. In the present format, up to 15 individuals can be typed per gel run. This method promises to provide the ability to develope large databases.
Subject Area
Genetics|Molecular biology
Recommended Citation
Jang, Gi Young, "A study of DNA identification using nonradioactive automated minisatellite variant repeat (MVR) digital typing" (1993). ETD collection for University of Nebraska-Lincoln. AAI9322801.
https://digitalcommons.unl.edu/dissertations/AAI9322801