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Y-STR analysis of degraded DNA using reduced-size amplicons

Abstract

To increase the success rate of Y-STR genotyping for degraded DNA, we have developed two multiplex PCR sets for 21 Y-STR loci. Besides the 17 Y-STR loci of DYS19, DYS385, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439, DYS448, DYS456, DYS458, DYS635, and GATA H4.1 contained in a commercial Y-STR kit, AmpFlSTR® Yfiler™, the other four loci of DYS388, DYS446, DYS447, and DYS449 were also included in the multiplexes to increase the discrimination capacity. Among a total of 21 Y-STR loci, the primers for eight loci (DYS385, DYS390, DYS438, DYS446, DYS448, DYS449, and DYS635) were newly designed in the present study and nine loci (DYS385, DYS390, DYS391, DYS392, DYS438, DYS439, DYS448, and DYS635) have PCR amplicons smaller than those of the AmpFlSTR® Yfiler™ kit. A sensitivity test using serially diluted standard 9948 male DNA showed that all the values of Y-STR loci in the Y-miniplexes are reliable at template concentrations as low as 30 pg. We compared the effectiveness of the two multiplexes with the AmpFlSTR® Yfiler™ kit by using both enzymatically degraded DNA and 30 samples of 50-year-old skeletal remains. This comparison demonstrated that the new Y-miniplex sets can produce a better signal from degraded DNA than the AmpFlSTR® Yfiler™ kit.

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References

  1. Roewer L, Arnemann J, Spurr NK, Grzeschik KH, Epplen JT (1992) Simple repeat sequences on the human Y chromosome are equally polymorphic as their autosomal counterparts. Hum Genet 89:389–394

    PubMed  Article  CAS  Google Scholar 

  2. Jobling MA, Tyler-Smith C (1995) Fathers and sons: the Y chromosome and human evolution. Trends Genet 11:449–456

    PubMed  Article  CAS  Google Scholar 

  3. Jobling MA, Pandya A, Tyler-Smith C (1997) The Y chromosome in forensic analysis and paternity testing. Int J Legal Med 110:118–124

    PubMed  Article  CAS  Google Scholar 

  4. Kayser M, Caglia A, Corach D et al (1997) Evaluation of Y-chromosomal STRs: a multicenter study. Int J Legal Med 110:125–133, 141–149

    PubMed  Article  CAS  Google Scholar 

  5. Berger B, Lindinger A, Niederstatter H, Grubwieser P, Parson W (2005) Y-STR typing of an Austrian population sample using a 17-loci multiplex PCR assay. Int J Legal Med 119:241–246

    PubMed  Article  Google Scholar 

  6. Kwak KD, Jin HJ, Shin DJ, Kim JM et al (2005) Y-chromosomal STR haplotypes and their applications to forensic and population studies in east Asia. Int J Legal Med 119:195–201

    PubMed  Article  Google Scholar 

  7. Bar W, Kratzer A, Machler M, Schmid W (1988) Postmortem stability of DNA. Forensic Sci Int 39:59–70

    PubMed  Article  CAS  Google Scholar 

  8. Alvarez Garcia A, Munoz I, Pestoni C, Lareu MV, Rodriguez-Calvo MS, Carracedo A (1996) Effect of environmental factors on PCR-DNA analysis from dental pulp. Int J Legal Med 109:125–129

    PubMed  Article  CAS  Google Scholar 

  9. Pfeiffer H, Huhne J, Seitz B, Brinkmann B (1999) Influence of soil storage and exposure period on DNA recovery from teeth. Int J Legal Med 112:142–144

    PubMed  Article  CAS  Google Scholar 

  10. Whitaker JP, Clayton TM, Urquhart AJ, Millican ES, Downes TJ, Kimpton CP, Gill P (1995) Short tandem repeat typing of bodies from a mass disaster: high success rate and characteristic amplification patterns in highly degraded samples. Biotechniques 18:670–677

    PubMed  CAS  Google Scholar 

  11. Schneider PM, Bender K, Mayr WR et al (2004) STR analysis of artificially degraded DNA—results of a collaborative European exercise. Forensic Sci Int 139:123–134

    PubMed  Article  CAS  Google Scholar 

  12. Yoshida K, Sekiguchi K, Kasai K, Sato H, Seta S, Sensabaugh GF (1997) Evaluation of new primers for CSF1PO. Int J Legal Med 110:36–38

    PubMed  CAS  Google Scholar 

  13. Hellmann A, Rohleder U, Schmitter H, Wittig M (2001) STR typing of human telogen hairs—a new approach. Int J Legal Med 114:269–273

    PubMed  Article  CAS  Google Scholar 

  14. Wiegand P, Kleiber M (2001) Less is more—length reduction of STR amplicons using redesigned primers. Int J Legal Med 114:285–287

    PubMed  Article  CAS  Google Scholar 

  15. Tsukada K, Takayanagi K, Asamura H, Ota M, Fukushima H (2002) Multiplex short tandem repeat typing in degraded samples using newly designed primers for the TH01, TPOX, CSF1PO, and vWA loci. Leg Med (Tokyo) 4:239–245

    CAS  Google Scholar 

  16. Butler JM, Shen Y, McCord BR (2003) The development of reduced size STR amplicons as tools for analysis of degraded DNA. J Forensic Sci 48:1054–1064

    PubMed  CAS  Google Scholar 

  17. Schilz F, Hummel S, Herrmann B (2004) Design of a multiplex PCR for genotyping 16 short tandem repeats in degraded DNA samples. Anthropol Anz 62:369–378

    PubMed  Google Scholar 

  18. Chung DT, Drabek J, Opel KL, Butler JM, McCord BR (2004) A study on the effects of degradation and template concentration on the amplification efficiency of the STR miniplex primer sets. J Forensic Sci 49:733–740

    PubMed  Article  CAS  Google Scholar 

  19. Coble MD, Butler JM (2005) Characterization of new miniSTR loci to aid analysis of degraded DNA. J Forensic Sci 50:43–53

    PubMed  Article  CAS  Google Scholar 

  20. Grubwieser P, Muhlmann R, Berger B, Niederstatter H, Pavlic M, Parson W (2005) A new “miniSTR-multiplex” displaying reduced amplicon lengths for the analysis of degraded DNA. Int J Legal Med 120:115–120

    PubMed  Article  Google Scholar 

  21. Wiegand P, Klein R, Braunschweiger G, Hohoff C, Brinkmann B (2006) Short amplicon STR multiplex for stain typing. Int J Legal Med 120:160–164

    PubMed  Article  CAS  Google Scholar 

  22. Roewer L, Epplen JT (1992) Rapid and sensitive typing of forensic stains by PCR amplification of polymorphic simple repeat sequences in case work. Forensic Sci Int 53:163–171

    PubMed  Article  CAS  Google Scholar 

  23. Schultes T, Hummel S, Herrmann B (1999) Amplification of Y-chromosomal STRs from ancient skeletal material. Hum Genet 104:164–166

    PubMed  Article  CAS  Google Scholar 

  24. Ayub Q, Mohyuddin A, Qamar R, Mazhar K, Zerjal T, Mehdi SQ, Tyler-Smith C (2000) Identification and characterisation of novel human Y-chromosomal microsatellites from sequence database information. Nucleic Acids Res 28:e8

    PubMed  Article  CAS  Google Scholar 

  25. Butler JM, Schoske R, Vallone PM, Kline MC, Redd AJ, Hammer MF (2002) A novel multiplex for simultaneous amplification of 20 Y chromosome STR markers. Forensic Sci Int 129:10–24

    PubMed  Article  CAS  Google Scholar 

  26. Redd AJ, Agellon AB, Kearney VA et al (2002) Forensic value of 14 novel STRs on the human Y chromosome. Forensic Sci Int 130:97–111

    PubMed  Article  CAS  Google Scholar 

  27. Schoske R, Vallone PM, Ruitberg CM, Butler JM (2003) Multiplex PCR design strategy used for the simultaneous amplification of 10 Y chromosome short tandem repeat (STR) loci. Anal Bioanal Chem 375:333–343

    PubMed  CAS  Google Scholar 

  28. Schoske R, Vallone PM, Kline MC, Redman JW, Butler JM (2004) High-throughput Y-STR typing of U.S. populations with 27 regions of the Y chromosome using two multiplex PCR assays. Forensic Sci Int 139:107–121

    PubMed  Article  CAS  Google Scholar 

  29. Vallone PM, Butler JM (2004) AutoDimer: a screening tool for primer-dimer and hairpin structures. Biotechniques 37:226–231

    PubMed  CAS  Google Scholar 

  30. Magnuson VL, Ally DS, Nylund SJ et al (1996) Substrate nucleotide-determined non-templated addition of adenine by Taq DNA polymerase: implications for PCR-based genotyping and cloning. Biotechniques 21:700–709

    PubMed  CAS  Google Scholar 

  31. Park MJ, Lee HY, Yoo JE, Chung U, Lee SY, Shin KJ (2005) Forensic evaluation and haplotypes of 19 Y-chromosomal STR loci in Koreans. Forensic Sci Int 152:133–147

    PubMed  Article  CAS  Google Scholar 

  32. Yang DY, Eng B, Waye JS, Dudar JC, Saunders SR (1998) Technical note: improved DNA extraction from ancient bones using silica-based spin columns. Am J Phys Anthropol 105:539–543

    PubMed  Article  CAS  Google Scholar 

  33. Gill P, Whitaker J, Flaxman C, Brown N, Buckleton J (2000) An investigation of the rigor of interpretation rules for STRs derived from less than 100 pg of DNA. Forensic Sci Int 112:17–40

    PubMed  Article  CAS  Google Scholar 

  34. Whitaker JP, Cotton EA, Gill P (2001) A comparison of the characteristics of profiles produced with the AMPFlSTR SGM Plus multiplex system for both standard and low copy number (LCN) STR DNA analysis. Forensic Sci Int 123:215–223

    PubMed  Article  CAS  Google Scholar 

  35. Taberlet P, Griffin S, Goossens B et al (1996) Reliable genotyping of samples with very low DNA quantities using PCR. Nucleic Acids Res 24:3189–3194

    PubMed  Article  CAS  Google Scholar 

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Acknowledgement

This work was supported by the Korea Research Foundation Grant funded by the Korean Government (MOEHRD, Basic Research Promotion Fund) (KRF-2004-003-E00004).

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Correspondence to Kyoung-Jin Shin.

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Table S1

Information on the 21 Y-STR loci of two multiplex PCR sets constructed in this study (PDF 89 kB)

Table S2

Primer sequences and concentrations of the optimized Y-miniplexes (PDF 68 kB)

Figure S1

Schematic of the PCR product size ranges produced with the known allele size ranges (see Table S1) for the loci in the two Y-miniplexes. Marker names were abbreviated (e.g. DYS391 is listed as 391)(PDF 77 kB)

Figure S2

Representative electropherograms of the allelic ladder and control 9948 male DNA amplified using the two Y-miniplexes.(PDF 150 kB)

Figure S3

Alignment of the top strands from DYS446 X and Y homologous sequences. The Y sequence comes from the GenBank accession number AC006152 while the X sequence comes from the GenBank accession number AL133512. Sequence difference between the X and Y homologs are boxed. The missing X sequence is indicated by dashes compared to the Y sequence. The primers indicated by dotted arrows are reported by Redd et al. [26] and these targeted a region containing four nucleotide differences between the X and Y sequences. The primers shown with solid arrows are the new ones described here that contains a single base difference in each of the forward and reverse primers (PDF 110 kB)

Figure S4

Sensitivity test for the two Y-miniplexes using control 9948 male DNAs in various concentrations: 1,000, 500, 250, 125, 62, 30 and 15 pg. The bar indicates the standard error from the average percent of alleles observed (PDF 109 kB)

Figure S5

Efficiency test using DNAs digested with DNase I for various time periods: 0, 2, 5, 10, 15, 20 and 30 min (PDF 606 kB)

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Park, M.J., Lee, H.Y., Chung, U. et al. Y-STR analysis of degraded DNA using reduced-size amplicons. Int J Legal Med 121, 152–157 (2007). https://doi.org/10.1007/s00414-006-0133-7

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  • DOI: https://doi.org/10.1007/s00414-006-0133-7

Keywords

  • Y chromosome
  • STR
  • AmpFlSTR Yfiler
  • Miniplex
  • Degraded DNA