Preface

1. Preface What follows is intended to be a brief review of the techniques employed by the ILE to collect, analyze and interpret DNA analysis results. We have not seen the Kercher’s case actual lab reports nor were we present when the samples were collected and successively stored, but we trust the judgment of several judges, and the reports of professional and well-reputed scientists working on the case who have stated international standard guidelines have always been followed in the stages of sample collection, storage, and DNA analysis.

2. DNA collection, profiling and interpretation: the ILE at work Content: -Suitable biological samples -Italian Forensic analysts: who are they, how do they collect samples, what DNA analysis protocols do they use? -Interpretation of Results

3. Step 1: collecting samples What are the biological samples more likely to be found on a crime scene? Human DNA can be extracted from any fluid or bodily (saliva, blood, semen, hair, nails etc) It can also be extracted from any object that has been contaminated by any bodily fluid such as chewing gum, cigarette butts, fabric, metals, paper tissues etc.

4. SAMPLES FROM WHICH DNA CAN BE EXTRACTED • blood • sperm • organic tissues • saliva • sweat • bones • teeth • urine • feces

5. Can DNA samples be contaminated during collection phase? Yes, they can. DNA can be contaminated by external sources, but since DNA DOES NOT FLY AND NEITHER IT WALKS, it can only be transferred passively. In order to understand the dynamics of passive DNA transfer, let’s consider two possible scenarios…

6. …Scenario no. 1 or THE INCOMPETENT COP Let’s say my neighbor gets murdered (God forbid!!). The cops knock at my door to ask me if I heard something. I let them in, and shake hands. The cats come by and one cat-lover cop stroke them. They enter my neighbor apartment and start collecting evidence wearing no gloves or no shoe-covers…. …They find traces of my DNA and cats hair on several pieces of evidence collected at the crime scene, including my neighbor bra and other items I had no business touching when paying friendly neighbor visits in the past. Although this wouldn’t rule me out as a suspect-unless of course I have a sound alibi-it could be a case of DNA contamination.

7. Scenario no. 2 or THE COMPETENT COP Let’s say my neighbor gets murdered (God forbid!!). The cops knock at my door to ask me if I heard something. I let them in, and shake hands. The cats come by and one cat-lover cop stroke them. They enter my neighbor apartment… …After they have carefully covered their shoes with shoe covers and wearing gloves that are changed every time they handle a new piece of evidence Do they find traces of my DNA and cats hair on some pieces of evidence collected at the crime scene? Perhaps, after all I was a frequent visitor there-assuming no bleaching has been performed-But do they find traces of my DNA on my neighbor bra or any other item I had no business to touch when paying friendly neighbor visits in the past? Very unlikely.

8. Unless someone can show the opposite occurred, we can say that correct sample collection techniques have been used at the crime scene in Via della Pergola. DNA is not a virus or a bacterium which can be transferred by breathing on objects. Contamination of the DNA specimens gathered at the murder scene was therefore extremely unlikely in the collection phase.

9. Who are the Scientific Police? • The Polizia Scientifica as well as the RIS –Reparto Investigazioni Scientifiche of the Carabinieri- are cops holding University and graduate degrees in biology, chemistry and physics, among others. • Many also hold a Master degree in Forensic Sciences. • The ILE operates according to the recommendations of the DNA Commission of the International Society for Forensic Genetics.

10. Storage of samples All samples (blood, saliva, tissues, objects etc) can be stored at room temperature for 24 hours. For longer periods the samples must be stored at – 20 C except for sample slides preparations, FABRICS AND OBJECTS that may continue to be stored at room temperature.

11. So, was the bra clasp contaminated or the DNA degraded in any way while sitting in sealed crime scene for a few weeks? Very unlikely

12. Analysis results interpretation according to the SIGU recommendations, following COUNCIL OF EUROPECommittee Recommendation No. R (92) 1 Genetic profiling results can lead to: Exclusion: the DNA cannot be traced back to the suspect Compatibility: only a partial analysis can be performed on “difficult substrates; identity cannot be ruled out but it is not attributable with high probability Identity: material present in discrete/good quality/quantity. Probability of having same identity extremely high.

13. PCR analysis • Contamination during this stage may always occur in any lab. However, several protocols exist that help to minimize the danger. • The biologist in charge has assured that maximum care was employed to avoid any chance of contamination. • However, test are always repeated just to make sure that results are not false positive (or even false negative)

14. Has the defendants’ DNA been found non-compatible with the traces found on the victim body/crime scene? NO Has the defendants’ DNA been found compatible with the traces found on the victim body/crime scene? NO Has the identity of the defendants’ been established as per their DNA found on the victim body/crime scene? YES

15. The analysis from 9 a 15 STR allows to establish a probability of match over 99,9999%, leading to a report of identity as signed by the biologist in charge of the analyses.

16. ENFSI CODIS (7 loci) (13 loci) TH01 TH01 D21S11 D21S11 D18S51 D18S51 vWA vWA FGA FGA D8S1179 D8S1179 TPOX CSF1PO D16S539 D7S820 D13S317 D5S818 D3S1358 D3S1358 STRs used in human DNA fingerprinting ENFSI (European Network of Forensic Science Institutes) CODIS (Combinated DNA Index System) della FBI.

17. Individuals differ from one another in terms of their STR profile. Currently, fifteen markers are being used in the determination of genetic profiles

18. Problems Given that during collection, handling and performing of the DNA analysis proper procedures have been followed as stated by several authoritative sources, sometimes interpretation problems may occur such as…

19. Unclear peaks • Unclear peaks due to technical problems; these peaks may be mistakenly interpreted as alleles

20. Possible Problems when DNA is scarce or degraded (Allele “dropout) Reference sample 1500 Sample to be analyzed 150 ? Sometimes only one of the two alleles at a locus is amplified. In the above figure, allele 14 at site D13S317 hasn’t amplified

21. However, it is well known how they are part of the most common defense strategies, such as : • Contamination • Statistical Weight of a Match • Degradation/PCR Inhibition of “True” Perp • Artifacts (N+4 stutter, etc.) • Thresholds Set Too High (missing peaks) • Examiner Bias • Improper Mixture Interpretation

22. We know at times some of it may happen… …however the evidence produced by prosecutors should be argued on the base of scientific expert knowledge and not by reasoning that… the crime took place in Italy hence the results-curiously limited to two of the defendants - must be wrong!

23. References • Hammond et al, 1994 Am J Hum Genet 55:175-189; • Garofano et al, 1998 J Forensic Sci 43: 837-840; • Garofano et al., 1998 Forensic Science Int. 97: 53-60) • www.bioforensic.com/conference07/index • John M. Butler "Forensic DNA typing: biology and technology behind STR markers".1st ed ACADEMIC PRESS 2001 • N. Rudin and K. Inman,“An introduction to Forensic DNA analysis”Second edition CRC Press 2002