DNA Sex Test: A New Rapid and Quantitative Forensic Approach Using Amelogenin Gene Based Fluorescent PCR

  • A. Mannucci
  • K. M. Sullivan
  • P. L. Ivanov
  • C. Kimpton
  • Peter Gill
Conference paper
Part of the Advances in Forensic Haemogenetics book series (HAEMOGENETICS, volume 5)

Abstract

For forensic PCR applications, it is recommended that the investigation of X and Y sequences should be carried out in parallel or simultaneously, and the distinction of male and female DNA cannot be made based solely on the absence of a band [1]. Several PCR-based tests have been developed for gender identification, among them multicopy repeat sequences on the X and Y chromosomes can be amplified together and provide a highly sensitive assay but quantitation of the relative X/Y product is not possible because of significant differences in repeat copy number [2–4]. Alternatively, single copy X-Y homologous regions such as amelogenin offer the advantage of requiring only one pair of primers and both X and Y sequences are of equal copy number [5]. Primers described by Sullivan and coll. flank a 6bp deletion within intron 1 of the X homologue resulting in 106bp and 112bp PCF, products from the X and Y chromosomes respectively [6]. Dye labelled PCR products were generated using one primer coupled with fluorescent dye ‘FAM’ via a 5’ aminolinker. These primers allowed DNA samples ranging from 10pg to 100ng to be amplified through 35 cycles comprising 1 minute at 94°C, 1 minute at 60°C and 1 minute at 72°C in a Perkin Elmer 480 thermal cycler. PCR products (20 μl) were visualised simply after electrophoresis in a 4% agarose gel for 90 mins at 100V and by ethidium bromide staining.

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References

  1. [1]
    Bar W., Brinkman B., Lincoln P., Mayr W.R., Rossi U. (1992) DNA recommendations-1992 report concerning recommendations of the DNA Commission of the International Society for Forensic Haemogenetics relating to the use of PCR-based polymorphisms. Int. J. Leg. Med., 105: 63–64.CrossRefGoogle Scholar
  2. [2]
    Witt M., Erickson R.P. (1989) A rapid method for the detection of Y-chromosomal DNA from dried blood specimens by the polymerase chain reaction. Hum. Genet. 82: 271–274.PubMedCrossRefGoogle Scholar
  3. [3]
    Pascal O., Aubert D., Gilbert E., Moisan J.P. (1993) Sexing of forensic samples using PCR. Int. J. Leg. Med. 104: 205–207.CrossRefGoogle Scholar
  4. [4]
    Gaensslen R.E., Berka K.M., Grosso D.A., Ruano G., Pagliaro EM, Messina D., Lee H.C. (1992) A polymerase chain reaction (PCR) method for sex and species determination with novel controls for deoxyribonucleic acid (DNA) template length. Journal of Forensic Sciences 37, 1: 6–20.PubMedGoogle Scholar
  5. [5]
    Nakahori Y., Hamano K., lwaya M., Nakagome Y. (1991) Sex identification by polymerase chain reaction using X-Y homologous primers. Am. J. Med. Genet. 39: 472–473.PubMedCrossRefGoogle Scholar
  6. [6]
    Sullivan K.M., Mannucci A., Kimpton C., Gill P. A Rapid and Quantitative DNA Sex Test: Fluorescence-based PCR analysis of X-Y homologous gene amelogenin“ submitted for publication in Biotechniques.Google Scholar
  7. [7]
    Sullivan K.M., A. Walton, C. Kimpton, G. Tully and P. Gill 1993 Fluorescence-based DNA segment analysis in forensic science, Biochemical Society Transactions 21: 1 16–120Google Scholar
  8. [8]
    Walsh, P.S., Erlich H.A., Higuchi R. (1992) Preferential PCR amplification of alleles: mechanisms and solutions. PCR Methods and Applications 1: 241–250.PubMedGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 1994

Authors and Affiliations

  • A. Mannucci
    • 1
  • K. M. Sullivan
  • P. L. Ivanov
  • C. Kimpton
  • Peter Gill
  1. 1.Istituto di Medicina LegaleUniversita’ di GenovaGenovaItaly

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