Amaç: Adli vakaların değerlendirilme sürecinde birden fazla kişinin DNA'sını içeren karışım örnekleri ile sıklıkla karşılaşılmaktadır. İki veya daha fazla kişinin genetik profilinin bir arada bulunduğu bu tür örneklerde, bazı faktörler (katılımcı sayısının bilinmemesi, allel çakışmalarının olması, düşük kalitede ve bozunmuş örnekler vb.) DNA analizini daha karmaşık hâle getirebilmektedir. Son yıllarda çeşitli ülkelerdeki hukuk sistemleri karışım DNA örneklerinin yorumlanmasında istatistiksel hesaplama işlemlerinden faydalanmaktadırlar. Ülkemizde karışım DNA analizlerinde kullanılacak herhangi bir prosedür bulunmamaktadır. Bu çalışmada; karışım örneklerinde profil olasılığı (GeneMapper® IDX v.1 Mixture Analysis) ve olabilirlik oranı (Forensim LRmix?) kullanılarak, hem doğru genotipleme yapmak hem de katılımcı sayısını doğru bir şekilde belirlemek amaçlanmıştır. Gereç ve Yöntemler: Bu çalışmada, gönüllü kişilerden toplanan kan örneklerinden değişik oranlarda (1:1, 1:3, 1:5, 1:10, 1:20, 1:50) karışım DNA örnekleri oluşturularak valide edilen AmpFℓSTR® Identifiler kiti ile çoğaltıldı. Polimeraz zincir reaksiyonu (PCR) ürünleri AB 3130 genetik analizörde yürütüldü ve Genemapper® IDX v.1 programında analiz edildi. Karışım örneklerini oluşturan katılımcıların tiplendirilmesi ve karışım oranlarının tahmin edilmesi Genemapper ® IDX v.1. Mixture Analysis ile yapıldı. Karışım örneklerinin allel düşmesi aralığına göre performans testi ve katılımcıların örnek profilinde bulunma olasılıklarının istatistiksel değerlendirmesi Forensim LRmix? uygulaması ile yapıldı. Bulgular: Analizde en sık karşılaşılan problem allel düşmesi olarak saptanmıştır. Olabilirlik oranı yaklaşımı ve Forensim LRmix? uygulaması kullanılarak oluşturulan karışım örneklerinin doğru bir şekilde DNA analizinin yapılabildiği belirlenmiştir. Sonuç: Karışım DNA örneklerinin objektif bir şekilde yorumlanabilmesi ve delilin gücünü ortaya koymak için istatistiksel yöntemler kullanılmalıdır. Bu çalışma; karışım DNA örneklerinin gelişmiş dünya ülkelerinde olduğu gibi ülkemizde de benzer standartlarda yorumlanabilmesi için yapılan öncü bir araştırmadır.
Anahtar Kelimeler: Adli genetik; kompleks DNA karışımları; mikrosatelit tekrarları; STR profili; LRmix modulu
Objective: In the process of evaluation of forensic cases, samples of the mixture containing more than one person are frequently encountered. In such examples of mixtures where the genetic profile of two or more persons are combined, some factors (Unknow number of contributors, presence of same alleles, poor quality and degraded samples, etc.) may further complicate DNA analysis. In recent years, legal systems in various countries have benefited from statistical calculations in the interpretation of mixture DNA samples. In our country, there are no procedures to be used in mixture DNA analysis. The aim of this study; to determine the correct genotypes and the number of contributors using the profile probability (GeneMapperR IDX v.1 Mixture Analysis) and likelihood ratio (Forensim LRmix?) in the mixture samples. Material and Methods: In this study, mixture samples were evaluated in different ratios (1:1, 1:3, 1:5, 1:10, 1:20, 1:50) from the blood samples which collected from volunteers. The polymerase chain reaction (PCR) was applied using validated AmpFℓSTRR Identifiler kit. PCR products were run in the AB 3130 enetic analyzer and analyzed in the GenemapperR IDX v.1 program. Typing of the contributors and estimating the mixture ratios were done on GenemapperR IDX v.1. Mixture Analysis. Forensim LRmix? application was used to test the performance of the mixture samples according to the allele drop interval and to evaluate the probabilities of the participants in the sample profile. Results: The most common problem in the analysis was the allele drop-out. Likelihood ratio and Forensim LRmix module was determined as a useful and successful application on both mixture samples that composed on controlled conditions. Conclusion: As a result, statistical methods should be used to interpret mixture DNA samples and to demonstrate the strength of evidence. This work as a pioneering research conducted in order to interpret mixed DNA samples in similar standards in our country as in developed countries.
Keywords: Forensic genetic; complex DNA mixtures; microsatellite repeats; Short tandem repeats; LRmix module
- Evett IW. Analysis of DNA multilocus profiles in a paternity case in which the child's profile may be partial. J Forensic Sci Soc. 1990;30(5):293-7. [Crossref ]
- Gill P, Sparkes R, Pinchin R, Clayton T, Whitaker J, Buckleton J. Interpreting simple STR mixtures using allele peak areas. Forensic Sci Int. 1998;91(1):41-53. [Crossref ]
- Butler JM. Short tandem repeat typing technologies used in human identity testing. Biotechniques. 2007;43(4):ii-v. [Crossref ]
- Gill P. The utility of 'substrate controls' in relation to 'contamination'. Forensic Sci Int. 1997;85(2):105-11. [Crossref ]
- Gill P, Brenner CH, Buckleton JS, Carracedo A, Krawczak M, Mayr WR, et al. DNA commission of the International Society of Forensic Genetics: Recommendations on the interpretation of mixtures. Forensic Sci Int. 2006;160(2-3):90-101. [Crossref ] [PubMed]
- Haned H, Pene L, Sauvage F, Pontier D. The predictive value of the maximum likelihood estimator of the number of contributors to a DNA mixture. Forensic Sci Int Genet. 2011;5(4): 281-4. [Crossref ] [PubMed]
- Haned H. Forensim: an open-source initiative for the evaluation of statistical methods in forensic genetics. Forensic Sci Int Genet. 2011;5(4):265-8. [Crossref ] [ PubMed]
- Swaminathan H, Qureshi MO, Grgicak CM, Duffy K, Lun DS. Four model variants within a continuous forensic DNA mixture interpretation framework: effects on evidential inference and reporting. PloS One. 2018;13(11): e0207599. [Crossref] [PubMed] [PMC]
- Bieber FR, Buckleton JS, Budowle B, Butler JM, Coble MD. Evaluation of forensic DNA mixture evidence: protocol for evaluation, interpretation, and statistical calculations using the combined probability of inclusion. BMC Genet. 2016;17(1):125. [Crossref ] [PubMed] [PMC]
- Budowle B, Onorato AJ, Callaghan TF, Della Manna A, Gross AM, Guerrieri RA, et al. Mixture interpretation: defining the relevant features for guidelines for the assessment of mixed DNA profiles in forensic casework. J Forensic Sci. 2009;54(4):810-21. [Crossref ] [PubMed]
- Haned H, Benschop CCG, Gill PD, Sijen T. Complex DNA mixture analysis in a forensic context: evaluating the probative value using a likelihood ratio model. Forensic Sci Int Genet. 2015;16:17-25. [Crossref ] [PubMed]
- Methods SWGoDA. SWGDAM Interpretation Guidelines for Autosomal STR Typing by Forensic DNA Testing Laboratories; 2017. p.28.
- Scientific TF. GeneMapper® ID-X Software Version 1.1 (Mixture Analysis Tool); 2008.
- Clayton D. On inferring presence of an individ ual in a mixture: a Bayesian approach. Biostatistics. 2010;11(4):661-73. [Crossref] [PubMed] [PMC]
- Collins PJ, Hennessy LK, Leibelt CS, Roby RK, Reeder DJ, Foxall PA. Developmental validation of a single-tube amplification of the 13 CODIS STR loci, D2S1338, D19S433, and amelogenin: the AmpFlSTR Identifiler PCR Amplification Kit. J Forensic Sci. 2004;49(6): 1265-77. [Crossref ] [PubMed]
- Malkoc E, Neuteboom W. The current status of forensic science laboratory accreditation in Europe. Forensic Sci Int. 2007;167(2-3):121-6. [Crossref] [PubMed]
- Szibor R, Edelmann J, Hering S, Plate I, Wittig H, Roewer L, et al. Cell line DNA typing in forensic genetics--the necessity of reliable standards. Forensic Sci Int. 2003;138(1-3):37-43. [Crossref ] [PubMed]
- Gill P, Brown RM, Fairley M, Lee L, Smyth M, Simpson N, et al. National recommendations of the Technical UK DNA working group on mixture interpretation for the NDNAD and for court going purposes. Forensic Sci Int Genet. 2008;2(1):76-82. [Crossref ] [PubMed]
- Bright JA, Turkington J, Buckleton J. Examination of the variability in mixed DNA profile parameters for the Identifiler multiplex. Forensic Sci Int Genet. 2010;4(2):111-4. [Crossref] [PubMed]
- Manabe S, Mori Y, Kawai C, Ozeki M, Tamaki K. Mixture interpretation: experimental and simulated reevaluation of qualitative analysis. Leg Med (Tokyo). 2013;15(2):66-71. [Crossref] [PubMed]
- Walsh PS, Fildes NJ, Reynolds R. Sequence analysis and characterization of stutter products at the tetranucleotide repeat locus vWA. Nucleic Acids Res. 1996;24(14):2807-12. [Crossref] [PubMed] [PMC]
- Leclair B, Frégeau CJ, Bowen KL, Fourney RM. Systematic analysis of stutter percentages and allele peak height and peak area ratios at heterozygous STR loci for forensic casework and database samples. J Forensic Sci. 2004;49(5):968-80. [Crossref ] [PubMed]
- Alonso A, Martín P, Albarrán C, García P, García O, de Simón LF, et al. Real-time PCR designs to estimate nuclear and mitochondrial DNA copy number in forensic and ancient DNA studies. Forensic Sci Int. 2004;139(2-3):141-9. [Crossref] [PubMed]
- Van Nieuwerburgh F, Goetghebeur E, Vandewoestyne M, Deforce D. Impact of allelic dropout on evidential value of forensic DNA profiles using RMNE. Bioinformatics. 2009;25(2):225-9. [Crossref ] [PubMed] [PMC]
- Tvedebrink T, Eriksen PS, Mogensen HS, Morling N. Estimating the probability of allelic dropout of STR alleles in forensic genetics. Forensic Sci Int Genet. 2009;3(3-4):222-6. [Crossref] [[PubMed]
- Gill P, Gusmão L, Haned H, Mayr WR, Morling N, Parson W, et al. DNA commission of the International Society of Forensic Genetics: recommendations on the evaluation of STR typing results that may include drop-out and/or dropin using probabilistic methods. Forensic Sci Int Genet. 2012;6(6):679-88. [Crossref ] [PubMed] [PMC]
- Prieto L, Haned H, Mosquera A, Crespillo M, Alemañ M, Aler M, et al. Euroforgen-NoE collaborative exercise on LRmix to demonstrate standardization of the interpretation of complex DNA profiles. Forensic Sci Int Genet. 2014;9:47-54. [Crossref][PubMed]
.: Process List