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“GenderPlex” a PCR multiplex for reliable gender determination of degraded human DNA samples and complex gender constellations

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Abstract

The amelogenin test integrated in most commercial polymerase chain reaction (PCR) multiplex kits is routinely used in the forensic field for gender determination of DNA samples. It has been demonstrated that this test is not entirely reliable. Males with deletions in the homologous amelogenin part on the Y chromosome (AMELY) were erroneously typed as females due to lack of Y-specific amelogenin amplification. Also, primer binding site mutations that result in a failure to amplify the AMELY or the X-chromosomal part (AMELX) have been observed. For clarification of such phenomena, a new PCR multiplex (GenderPlex) is presented, co-amplifying two different regions of the amelogenin gene (55/58 and 106/112 bp for the AMELX and AMELY alleles, respectively), a 93-bp sequence stretch of the SRY gene and four mini-X-STR loci DXS7424, DXS8378, DXS6803 and GATA172D05 (maximum product size less than 140 bp). This strategy helps with the evaluation of samples for the presence of amelogenin-based primer site mutations and confirms a male genotype by the absence of heterozygote X-STR alleles and the presence of an SRY-related peak. The short amplicon sizes of all involved loci proved to be beneficial in a study on artificially degraded DNA. Furthermore, we demonstrate by means of sensitivity, human specificity and mixture studies that the multiplex is suitable for investigations in the forensic scene. Finally, the performance of the GenderPlex was evaluated on a west Eurasian population sample from Austria comprising 166 male and 104 female individuals.

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Acknowledgements

The authors would like to thank Volker Büchele, the Tiroler Landesmuseum Ferdinandeum, the Naturwissenschaftliche Sammlungen “Alpenzoo Innsbruck” and the “Tierpark Hellabrunn” in Munich for kindly providing non-human samples. Cordula Berger and Bettina Zimmermann are greatly acknowledged for their helpful assistance, comments and discussion.

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Authors and Affiliations

  1. Institute of Legal Medicine, Innsbruck Medical University, Müllerstrasse 44, 6020, Innsbruck, Austria

    Anna Esteve Codina, Harald Niederstätter & Walther Parson

Authors
  1. Anna Esteve Codina
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  2. Harald Niederstätter
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  3. Walther Parson
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Corresponding author

Correspondence to Walther Parson.

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Table S1

Primers for sequence analysis of the X-STR ladder alleles (DOC 29 KB)

Table S2

Apparent GenderPlex PCR product lengths (nucleotides) obtained for different vertebrate species (DOC 112 KB)

Table S3

Relative allele frequencies of the X-STRs in the west Eurasian population sample from Austria (166 male and 104 female donors) (DOC 45.5 KB)

Table S4

Genotypes and their relative frequencies found in the male west Eurasian population sample from Austria (N = 166) (DOC 336 KB)

Table S5

Genotypes of the female west Eurasian population sample from Austria (N = 104) (DOC 339 KB)

Figure S1

GenderPlex electropherogram from a male person lacking AMELY but displaying full minHT Y-STR [8], SRY and single X-STRs peaks (PPT 54.5 KB)

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Esteve Codina, A., Niederstätter, H. & Parson, W. “GenderPlex” a PCR multiplex for reliable gender determination of degraded human DNA samples and complex gender constellations. Int J Legal Med 123, 459–464 (2009). https://doi.org/10.1007/s00414-008-0301-z

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  • DOI: https://doi.org/10.1007/s00414-008-0301-z

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