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Advances in the DNA analysis of canine trace evidence for serious crime investigation in the UK
Forensic Science International: Genetics Supplement Series 2 (2009) 290–291
Contents lists available at ScienceDirect
Forensic Science International: Genetics Supplement Series journal homepage: www.elsevier.com/locate/FSIGSS
Research article
Advances in the DNA analysis of canine trace evidence for serious crime investigation in the UK Rob Ogden a,*, Elizabeth Heap b, Ross McEwing c a
Gen-Probe Life Sciences, Appleton Place, Livingston EH54 7EZ, United Kingdom The School of Biological Sciences, University of Edinburgh, King’s Buildings, Mayfield Road, Edinburgh EH9 3JR, United Kingdom c TRACE Wildlife Forensics Network, Royal Zoological Society of Scotland, Edinburgh EH12 6TS, United Kingdom b
A R T I C L E I N F O
A B S T R A C T
Article history: Received 14 September 2009 Accepted 16 September 2009
Non-human evidence is becoming widespread in serious crime investigations. Canine hair samples have proved especially useful due to the high rate of transfer that naturally occurs between dogs, people and property. A murder investigation in the UK during 2008 provided the opportunity to undertake further research into the application and limitations of non-human hairs for linking victim to suspect. The overall aim of the work was to maximize the genetic data that could be obtained from shed hairs. This was attempted by conducting four short experiments to examine: (i) the recovery of exogenous human DNA from the surface of hairs, (ii) the potential for canine DNA originating from saliva to be obtained from hair shafts, (iii) the presence of residual DNA on tape lifts following hair removal and (iv) the potential for enhanced profile recovery following whole genome amplification of canine DNA. Results showed that DNA could be recovered from the surface of hairs following simulated licking and skin contact. When applied to casework samples, additional human DNA evidence was recovered from dog hairs in this manner. Attempted DNA recovery from tape lifts was unsuccessful. Whole genome amplification improved the recovery of full and partial profiles under reducing DNA concentration. The work demonstrates several novel approaches for recovering trace genetic evidence from animal hairs. Although such detailed analysis is unlikely to be applied to routine casework investigations, it increases the range of forensic tools available to investigate serious crimes. ß 2009 Elsevier Ireland Ltd. All rights reserved.
Keywords: Non-human forensics mtDNA Animal hairs Trace evidence Dog Forensic
1. Introduction The use of non-human evidence is becoming widespread in serious crime investigation. Canine samples have proved particularly useful due to the high rate of trace evidence transfer from dogs to humans. Dog hairs are routinely used as a source of canine DNA, however in the absence of root cells analysis is usually restricted to mtDNA within the hair shaft. A murder investigation in the UK in 2008 yielded a large number of suspected canine hair fragments thought to link the suspect to the victim via the suspect’s dog. To maximize the recovery of DNA evidence from dog hairs, research was undertaken to assess four novel analytical methods. 1.1. Aims To examine the recovery of human contact DNA from the exterior surface of dog hairs.
* Corresponding author. Tel.: +44 1506 424290; fax: +44 1506 424280. E-mail address: [email protected] (R. Ogden). 1875-1768/$ – see front matter ß 2009 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.fsigss.2009.09.033
To examine the recovery of canine DNA from saliva deposited on hair shafts during grooming. To examine the recovery of canine DNA from tape lift residues, following the removal of canine hairs. To assess the use of whole genome amplification in the recovery of DNA STR profile data from trace dog samples. 2. Materials and methods 2.1. The recovery of human contact DNA from dog hairs This experiment was designed to assess whether or not human DNA could be recovered from the shaft of individual dog hairs resulting from routine contact (patting/stroking etc.). Five individual dog hairs (4 cm long) were handled by five technical staff, one hair per person. Each hair was rubbed repeatedly in the palm of the hand and fingers for 15 s. Hair samples were submitted to LGC Forensics for DNA profiling using the SGM+ marker panel in combination with the standard 28-cycle analysis and the advanced SenCE 62 analysis. Results were compared with control samples (buccal swabs) from the same individuals.
R. Ogden et al. / Forensic Science International: Genetics Supplement Series 2 (2009) 290–291
291
2.2. The recovery of canine DNA from saliva deposited onto hair shafts
3.2. The recovery of canine DNA from saliva deposited onto hair shafts
This experiment aimed to examine the possibility that nuclear DNA could be recovered from canine saliva transferred to the outside of hair shafts during self-grooming/licking. Dog saliva was obtained from a reference dog and applied to individual cat hairs and allowed to dry. Samples were stored for up to two weeks before DNA was recovered by washing individual hairs and extracting from the wash solution. Each sample was then submitted for mtDNA sequence and nDNA STR profile analysis.
Canine mitochondrial DNA sequences and partial nuclear DNA profiles matching the contributor dog were obtained from hair washes after two weeks. No cat DNA was amplified from the hair wash extract using universal mammalian primers. These results demonstrate the transfer of canine DNA from saliva to hair shaft and importantly raise the possibility of recovering nuclear DNA from hair samples via external deposition. 3.3. The recovery of DNA from tape lifts, following the removal of hairs
2.3. The recovery of DNA from tape lifts, following the removal of hairs Following the results of experiment 2, it was considered possible that exogenous hair shaft DNA would remain on plastic tape lifts after removal of hairs for processing. The purpose of this experiment was to examine the feasibility of analyzing any such DNA. Four tape lifts were prepared containing cat hairs coated with dried dog saliva. The hairs were first removed from the tape lifts and submitted for analysis. Next, thin strips of the plastic tape lift were dissected from where the hairs had been attached and submitted for comparison. 2.4. The use of whole genome amplification to enhance DNA analysis This experiment was conducted to examine whether or not the recovery of DNA profiles was improved with the addition of a whole genome amplification (WGA) step after DNA extraction and directly before marker-specific PCR. Four canine DNA samples were analysed using eight STR markers across serial dilutions of template DNA (0.9 ng–0.9 pg), with and without the use of the WGA using the Qiagen REPLI-g kit. 3. Results and discussion 3.1. The recovery of human contact DNA from dog hairs For four of the five samples, no human STR profile information was recovered with the exception of single observations for two alleles. For the fifth sample, a partial profile was recovered under both sets of profiling conditions which matched the profile of the contributor. This result provides some support for the proposition of recovering human contact DNA from dog hairs, however the success rate is likely to be extremely low for casework samples and this approach would therefore probably be restricted to use where little alternative evidence was available.
No amplifiable DNA was recovered from the areas of plastic taping that had been in contact with the hairs. The hairs themselves were successfully analysed for feline DNA. In combination with the saliva results described above, the data indicate that exogenous hair shaft DNA is not readily transferred and retained on tape lifts. 3.4. The use of whole genome amplification to enhance DNA analysis Results for the experiment showed that enhanced profile information was recovered for samples that have undergone WGA (Fig. 1), particularly for very low starting template DNA amounts. The percentage recovery of alleles across all STR loci was markedly improved at 0.9 and 9.0 pg template DNA following WGA, enabling partial profiles to be observed where none were produced under standard amplification alone. At higher template DNA amounts, WGA-treated samples displayed less allelic drop-out than the untreated controls and, importantly, were always consistent with the reference sample control profile. The findings of the four experiments extend the range of forensic genetic tools available for canine hair samples and raise the potential for new evidence to be obtained capable of identifying both individual dogs and humans. Some of the methods described are already being used in the UK and for serious crimes with sufficient analytical resources such techniques may play an important part in future forensic investigation. 4. Conclusions 1. Canine hairs may provide a source of transfer human DNA. 2. Canine nDNA and mtDNA can be recovered from saliva residues on single hairs. 3. There was no evidence that exogenous DNA on hairs is transferred to tape lifts. 4. Whole genome amplification can improve the recovery of canine STR profiles. Conflict of interest None. Role of funding statement The research reported here was funded privately as research to support an ongoing casework investigation. The sponsor was involved in one stage of the genetic analysis (human STR profiling) but had no role in study design; in the non-human DNA analysis; in the interpretation of data; in the writing of the report; nor in the decision to submit the paper for publication. Acknowledgment
Fig. 1. A comparison of STR profiling methods with and without a whole genome amplification (WGA) stage prior to PCR. The recovery of target alleles is much higher in WGA genomic DNA (WGA gDNA) than in the non-WGA treatment at low DNA template amounts, supporting the use of WGA for low yield sample types.
The authors would like to thank LGC Forensics for their contribution to the laboratory work presented here.
Contents lists available at ScienceDirect
Forensic Science International: Genetics Supplement Series journal homepage: www.elsevier.com/locate/FSIGSS
Research article
Advances in the DNA analysis of canine trace evidence for serious crime investigation in the UK Rob Ogden a,*, Elizabeth Heap b, Ross McEwing c a
Gen-Probe Life Sciences, Appleton Place, Livingston EH54 7EZ, United Kingdom The School of Biological Sciences, University of Edinburgh, King’s Buildings, Mayfield Road, Edinburgh EH9 3JR, United Kingdom c TRACE Wildlife Forensics Network, Royal Zoological Society of Scotland, Edinburgh EH12 6TS, United Kingdom b
A R T I C L E I N F O
A B S T R A C T
Article history: Received 14 September 2009 Accepted 16 September 2009
Non-human evidence is becoming widespread in serious crime investigations. Canine hair samples have proved especially useful due to the high rate of transfer that naturally occurs between dogs, people and property. A murder investigation in the UK during 2008 provided the opportunity to undertake further research into the application and limitations of non-human hairs for linking victim to suspect. The overall aim of the work was to maximize the genetic data that could be obtained from shed hairs. This was attempted by conducting four short experiments to examine: (i) the recovery of exogenous human DNA from the surface of hairs, (ii) the potential for canine DNA originating from saliva to be obtained from hair shafts, (iii) the presence of residual DNA on tape lifts following hair removal and (iv) the potential for enhanced profile recovery following whole genome amplification of canine DNA. Results showed that DNA could be recovered from the surface of hairs following simulated licking and skin contact. When applied to casework samples, additional human DNA evidence was recovered from dog hairs in this manner. Attempted DNA recovery from tape lifts was unsuccessful. Whole genome amplification improved the recovery of full and partial profiles under reducing DNA concentration. The work demonstrates several novel approaches for recovering trace genetic evidence from animal hairs. Although such detailed analysis is unlikely to be applied to routine casework investigations, it increases the range of forensic tools available to investigate serious crimes. ß 2009 Elsevier Ireland Ltd. All rights reserved.
Keywords: Non-human forensics mtDNA Animal hairs Trace evidence Dog Forensic
1. Introduction The use of non-human evidence is becoming widespread in serious crime investigation. Canine samples have proved particularly useful due to the high rate of trace evidence transfer from dogs to humans. Dog hairs are routinely used as a source of canine DNA, however in the absence of root cells analysis is usually restricted to mtDNA within the hair shaft. A murder investigation in the UK in 2008 yielded a large number of suspected canine hair fragments thought to link the suspect to the victim via the suspect’s dog. To maximize the recovery of DNA evidence from dog hairs, research was undertaken to assess four novel analytical methods. 1.1. Aims To examine the recovery of human contact DNA from the exterior surface of dog hairs.
* Corresponding author. Tel.: +44 1506 424290; fax: +44 1506 424280. E-mail address: [email protected] (R. Ogden). 1875-1768/$ – see front matter ß 2009 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.fsigss.2009.09.033
To examine the recovery of canine DNA from saliva deposited on hair shafts during grooming. To examine the recovery of canine DNA from tape lift residues, following the removal of canine hairs. To assess the use of whole genome amplification in the recovery of DNA STR profile data from trace dog samples. 2. Materials and methods 2.1. The recovery of human contact DNA from dog hairs This experiment was designed to assess whether or not human DNA could be recovered from the shaft of individual dog hairs resulting from routine contact (patting/stroking etc.). Five individual dog hairs (4 cm long) were handled by five technical staff, one hair per person. Each hair was rubbed repeatedly in the palm of the hand and fingers for 15 s. Hair samples were submitted to LGC Forensics for DNA profiling using the SGM+ marker panel in combination with the standard 28-cycle analysis and the advanced SenCE 62 analysis. Results were compared with control samples (buccal swabs) from the same individuals.
R. Ogden et al. / Forensic Science International: Genetics Supplement Series 2 (2009) 290–291
291
2.2. The recovery of canine DNA from saliva deposited onto hair shafts
3.2. The recovery of canine DNA from saliva deposited onto hair shafts
This experiment aimed to examine the possibility that nuclear DNA could be recovered from canine saliva transferred to the outside of hair shafts during self-grooming/licking. Dog saliva was obtained from a reference dog and applied to individual cat hairs and allowed to dry. Samples were stored for up to two weeks before DNA was recovered by washing individual hairs and extracting from the wash solution. Each sample was then submitted for mtDNA sequence and nDNA STR profile analysis.
Canine mitochondrial DNA sequences and partial nuclear DNA profiles matching the contributor dog were obtained from hair washes after two weeks. No cat DNA was amplified from the hair wash extract using universal mammalian primers. These results demonstrate the transfer of canine DNA from saliva to hair shaft and importantly raise the possibility of recovering nuclear DNA from hair samples via external deposition. 3.3. The recovery of DNA from tape lifts, following the removal of hairs
2.3. The recovery of DNA from tape lifts, following the removal of hairs Following the results of experiment 2, it was considered possible that exogenous hair shaft DNA would remain on plastic tape lifts after removal of hairs for processing. The purpose of this experiment was to examine the feasibility of analyzing any such DNA. Four tape lifts were prepared containing cat hairs coated with dried dog saliva. The hairs were first removed from the tape lifts and submitted for analysis. Next, thin strips of the plastic tape lift were dissected from where the hairs had been attached and submitted for comparison. 2.4. The use of whole genome amplification to enhance DNA analysis This experiment was conducted to examine whether or not the recovery of DNA profiles was improved with the addition of a whole genome amplification (WGA) step after DNA extraction and directly before marker-specific PCR. Four canine DNA samples were analysed using eight STR markers across serial dilutions of template DNA (0.9 ng–0.9 pg), with and without the use of the WGA using the Qiagen REPLI-g kit. 3. Results and discussion 3.1. The recovery of human contact DNA from dog hairs For four of the five samples, no human STR profile information was recovered with the exception of single observations for two alleles. For the fifth sample, a partial profile was recovered under both sets of profiling conditions which matched the profile of the contributor. This result provides some support for the proposition of recovering human contact DNA from dog hairs, however the success rate is likely to be extremely low for casework samples and this approach would therefore probably be restricted to use where little alternative evidence was available.
No amplifiable DNA was recovered from the areas of plastic taping that had been in contact with the hairs. The hairs themselves were successfully analysed for feline DNA. In combination with the saliva results described above, the data indicate that exogenous hair shaft DNA is not readily transferred and retained on tape lifts. 3.4. The use of whole genome amplification to enhance DNA analysis Results for the experiment showed that enhanced profile information was recovered for samples that have undergone WGA (Fig. 1), particularly for very low starting template DNA amounts. The percentage recovery of alleles across all STR loci was markedly improved at 0.9 and 9.0 pg template DNA following WGA, enabling partial profiles to be observed where none were produced under standard amplification alone. At higher template DNA amounts, WGA-treated samples displayed less allelic drop-out than the untreated controls and, importantly, were always consistent with the reference sample control profile. The findings of the four experiments extend the range of forensic genetic tools available for canine hair samples and raise the potential for new evidence to be obtained capable of identifying both individual dogs and humans. Some of the methods described are already being used in the UK and for serious crimes with sufficient analytical resources such techniques may play an important part in future forensic investigation. 4. Conclusions 1. Canine hairs may provide a source of transfer human DNA. 2. Canine nDNA and mtDNA can be recovered from saliva residues on single hairs. 3. There was no evidence that exogenous DNA on hairs is transferred to tape lifts. 4. Whole genome amplification can improve the recovery of canine STR profiles. Conflict of interest None. Role of funding statement The research reported here was funded privately as research to support an ongoing casework investigation. The sponsor was involved in one stage of the genetic analysis (human STR profiling) but had no role in study design; in the non-human DNA analysis; in the interpretation of data; in the writing of the report; nor in the decision to submit the paper for publication. Acknowledgment
Fig. 1. A comparison of STR profiling methods with and without a whole genome amplification (WGA) stage prior to PCR. The recovery of target alleles is much higher in WGA genomic DNA (WGA gDNA) than in the non-WGA treatment at low DNA template amounts, supporting the use of WGA for low yield sample types.
The authors would like to thank LGC Forensics for their contribution to the laboratory work presented here.