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Published in: International Journal of Legal Medicine 6/2003

01-12-2003 | Original Article

An easily automated, closed-tube forensic DNA extraction procedure using a thermostable proteinase

Authors: D. Moss, S.-A. Harbison, D. J. Saul

Published in: International Journal of Legal Medicine | Issue 6/2003

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Abstract

Most standard procedures for extracting DNA from forensic substrates involve manipulations that expose the sample to potential contamination and which reduce yields. Furthermore, most methods require centrifugation and/or solvent extraction steps that render them difficult to automate. We describe a simple closed-tube DNA extraction procedure using a proteinase from the thermophilic Bacillus species EA1 that produces good DNA yields from a wide range of forensic substrates. The reaction is controlled by a temperature shift regime programmed into a thermal cycler and so eliminates the need for solvent extraction or column purification. The new method is ideally suited to forensic samples where exposure to extraneous contaminating DNA must be avoided. In addition, The simplicity of the procedure makes it suitable for automation.
Literature
1.
go back to reference McHale R, Stapleton PM, Bergquist PL (1991) A rapid method for the preparation of samples for PCR. Biotechniques 10:20–22PubMed McHale R, Stapleton PM, Bergquist PL (1991) A rapid method for the preparation of samples for PCR. Biotechniques 10:20–22PubMed
2.
go back to reference Borges KM, Bergquist PL (1993) Pulsed-field gel electrophoresis study of the genome of Caldocellum saccharolyticum. Curr Microbiol 27:15–19 Borges KM, Bergquist PL (1993) Pulsed-field gel electrophoresis study of the genome of Caldocellum saccharolyticum. Curr Microbiol 27:15–19
3.
go back to reference Collet C (2001) Proteases suitable for use in molecular biology. Master of philosophy thesis, University of Waikato, Hamilton, NZ Collet C (2001) Proteases suitable for use in molecular biology. Master of philosophy thesis, University of Waikato, Hamilton, NZ
4.
go back to reference Coolbear T, Eames CW, Casey Y, Daniel RM, Morgan HW (1991) Screening of strains identified as extremely thermophilic bacilli for extracellular proteolytic activity and general properties of the proteinases from two of the isolates. J Appl Bacteriol 71:252–264 Coolbear T, Eames CW, Casey Y, Daniel RM, Morgan HW (1991) Screening of strains identified as extremely thermophilic bacilli for extracellular proteolytic activity and general properties of the proteinases from two of the isolates. J Appl Bacteriol 71:252–264
5.
go back to reference Coolbear T, Whittaker JM, Daniel RM, Morgan HW (1992) The effect of metal ions on the activity and thermostability of the extracellular proteinase from a thermophilic Bacillus strain EA1. Biochem J 286:367–374 Coolbear T, Whittaker JM, Daniel RM, Morgan HW (1992) The effect of metal ions on the activity and thermostability of the extracellular proteinase from a thermophilic Bacillus strain EA1. Biochem J 286:367–374
6.
go back to reference Saul DJ, Williams LC, Toogood HS, Daniel RM, Bergquist PL (1996) Sequence of the gene encoding a highly thermostable neutral proteinase from Bacillus sp. strain EA1: expression in Escherichia coli and characterisation. Biochim Biophys Acta 1308:74–80CrossRefPubMed Saul DJ, Williams LC, Toogood HS, Daniel RM, Bergquist PL (1996) Sequence of the gene encoding a highly thermostable neutral proteinase from Bacillus sp. strain EA1: expression in Escherichia coli and characterisation. Biochim Biophys Acta 1308:74–80CrossRefPubMed
7.
go back to reference Linch C, Smith SL, Prahlow JA (1998) Evaluation of the human hair root for DNA typing subsequent to microscopic comparison. J Forensic Sci 43:305–314PubMed Linch C, Smith SL, Prahlow JA (1998) Evaluation of the human hair root for DNA typing subsequent to microscopic comparison. J Forensic Sci 43:305–314PubMed
8.
go back to reference Sweet D, Lorente M, Lorente J, Valenzuela A, Villanueva E (1997) An improved method to recover saliva from human skin: the double swab technique. J Forensic Sci 42:320–322PubMed Sweet D, Lorente M, Lorente J, Valenzuela A, Villanueva E (1997) An improved method to recover saliva from human skin: the double swab technique. J Forensic Sci 42:320–322PubMed
9.
go back to reference Walsh PS, Metzge D, Higuchi R (1991) Chelex 100 as a medium for simple extraction of DNA for PCR-based typing from forensic material. Biotechniques 10:506–513PubMed Walsh PS, Metzge D, Higuchi R (1991) Chelex 100 as a medium for simple extraction of DNA for PCR-based typing from forensic material. Biotechniques 10:506–513PubMed
10.
go back to reference Sambrook J, Fritsch DW (2001) Molecular cloning, a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, New York, USA Sambrook J, Fritsch DW (2001) Molecular cloning, a laboratory manual, 3rd edn. Cold Spring Harbor Laboratory Press, New York, USA
11.
go back to reference Moretti T, Baumstark AL, Defenbaugh BS, Keys KM, Smerick JB, Budowle B (2001) Validation of Short Tandem Repeats (STRs) for forensic usage: performance testing of fluorescent multiplex STR systems and analysis of authentic and simulated forensic samples. J Forensic Sci 3:647–660 Moretti T, Baumstark AL, Defenbaugh BS, Keys KM, Smerick JB, Budowle B (2001) Validation of Short Tandem Repeats (STRs) for forensic usage: performance testing of fluorescent multiplex STR systems and analysis of authentic and simulated forensic samples. J Forensic Sci 3:647–660
12.
go back to reference Gill P, Sparkes RL, Pinchin R, Clayton T, Whitaker JP, Buckleton J (1998) Interpreting simple STR mixtures using allelic peak areas. Forensic Sci Int 91:41–53CrossRefPubMed Gill P, Sparkes RL, Pinchin R, Clayton T, Whitaker JP, Buckleton J (1998) Interpreting simple STR mixtures using allelic peak areas. Forensic Sci Int 91:41–53CrossRefPubMed
13.
go back to reference Clayton T, Whitaker JP, Sparkes RL, Gill P (1998) Analysis and interpretation of mixed forensic stains using DNA STR profiling. Forensic Sci Int 91:55–70CrossRefPubMed Clayton T, Whitaker JP, Sparkes RL, Gill P (1998) Analysis and interpretation of mixed forensic stains using DNA STR profiling. Forensic Sci Int 91:55–70CrossRefPubMed
14.
go back to reference Akane A, Matsubara K, Nakamura H, Takahashi S, Kimura K (1994) Identification of the heme compound co-purified with deoxyribonucleic acid (DNA) from bloodstains. A major inhibitor of the polymerase chain reaction (PCR) amplification. J Forensic Sci 39:362–372PubMed Akane A, Matsubara K, Nakamura H, Takahashi S, Kimura K (1994) Identification of the heme compound co-purified with deoxyribonucleic acid (DNA) from bloodstains. A major inhibitor of the polymerase chain reaction (PCR) amplification. J Forensic Sci 39:362–372PubMed
15.
go back to reference Klintschar M, Neuhuber F (2000) Evaluation of an alkaline lysis method for the extraction of DNA from whole blood and forensic stains for STR analysis. J Forensic Sci 45:669–673PubMed Klintschar M, Neuhuber F (2000) Evaluation of an alkaline lysis method for the extraction of DNA from whole blood and forensic stains for STR analysis. J Forensic Sci 45:669–673PubMed
16.
go back to reference Matsuda H, Imaizumi K, Kubota S, Miyasaka S, Yoshino M, Seta S (1997) Technical investigation of DNA extraction from a single hair shaft. Rep Nat Res Inst Police Sci 50:23–28 Matsuda H, Imaizumi K, Kubota S, Miyasaka S, Yoshino M, Seta S (1997) Technical investigation of DNA extraction from a single hair shaft. Rep Nat Res Inst Police Sci 50:23–28
17.
go back to reference Van Oorschot RAH, Phelan DG, Furlong S, Scarfo GM, Holding NL, Cummins MJ (2003) Are you collecting all the available DNA from touched objects? In: Brinkmann B, Carracedo A (eds) Progress in forensic genetics 9. Elsevier, Amsterdam, pp 803–807 Van Oorschot RAH, Phelan DG, Furlong S, Scarfo GM, Holding NL, Cummins MJ (2003) Are you collecting all the available DNA from touched objects? In: Brinkmann B, Carracedo A (eds) Progress in forensic genetics 9. Elsevier, Amsterdam, pp 803–807
18.
go back to reference Jehaes E, Toprak K, Vanderheyen H, Pfeiffer H, Brinkmann B, Decorte R (2001) Pitfalls in the analysis of mitochondrial DNA from ancient specimens and the consequences for forensic analysis: the historical case of the putative heart of Louis XVII. Int J Legal Med 115:135–141PubMed Jehaes E, Toprak K, Vanderheyen H, Pfeiffer H, Brinkmann B, Decorte R (2001) Pitfalls in the analysis of mitochondrial DNA from ancient specimens and the consequences for forensic analysis: the historical case of the putative heart of Louis XVII. Int J Legal Med 115:135–141PubMed
19.
go back to reference Hellmann A, Rohleder U, Schmitter H, Wittig M (2001) STR typing of human telogen hairs—a new approach. Int J Legal Med 114:269–273PubMed Hellmann A, Rohleder U, Schmitter H, Wittig M (2001) STR typing of human telogen hairs—a new approach. Int J Legal Med 114:269–273PubMed
Metadata
Title
An easily automated, closed-tube forensic DNA extraction procedure using a thermostable proteinase
Authors
D. Moss
S.-A. Harbison
D. J. Saul
Publication date
01-12-2003
Publisher
Springer-Verlag
Published in
International Journal of Legal Medicine / Issue 6/2003
Print ISSN: 0937-9827
Electronic ISSN: 1437-1596
DOI
https://doi.org/10.1007/s00414-003-0400-9

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