1. 3 Person example #1 Victim Vaginal Swab Victim and Consensual Partner Who Did It

2. The Scenario Victim hosted a party at her place She had sex with her boyfriend prior to guests arriving She drank too much and went bed before all guests left She dreamed that someone had sex with her – boyfriend was still there

3. The Scenario She woke up and IDd Suspect leaving her room Suspect admitted he was in the room, but it was just to grab an X-box game he had left in there

4. The Egram

5. Sample Interpretation Consistent with 3 persons Appears to have high- middle- and low-level contributors Perhaps the low level contributor drops out? – Some minor alleles <300 – Such as our good friend FGA X and Y somewhat balanced – Will we be lucky and have Victim as the minor?

6. The Data Table

7. Match to Victim Victim fits every where but FGA She has a 22 there that is missing from the mixture She is the low-level contributor

8. Match to Consensual All alleles of Consensual present He is 20, 23 at D2 (6 Alleles) So he is high-level contributor (State assumption in report)

9. Match to Suspect All alleles of Suspect are found

10. How many would assume 1.Victim only 2.Consensual only 3.Victim and Consensual 4.No one

11. My approach I assume both V and C, and will communicate that in the report Because V is minor, Im not concerned about alleles <300, as I know who they belong to I have a pretty good idea that C is the major (or at least the majorish) Im interested in the in-between contributor

12. My approach I know we already did the match to the Suspect, but well hide his profile before we start looking at the mixture I already decided the mixture is interpretable as a three person mixture, and I have lots of ways to deal with the stat

13. Any conclusions about Suspect? 1.Match 2.Included 3.Inconclusive 4.Excluded 5.None of the above - Profile cannot be interpreted

14. The Deconvolution Well hide the Suspects type from the table – No peeking! Remember to put the calculator into 3 person mode (Not a trivial step) Lets go highest to lowest in terms of number of alleles in our interpretation Well apply both Victim and Consensual and try to deconvolute Unknown type I have to do those 4 things now

15. The Egram

16. The Deconvolution Start with D2 (six alleles) – And nothing fits!

17. The Deconvolution Recalculate for stutter at D2 Still didnt help – but Im smarter than the software so Im over ruling it

18. The Deconvolution Why am I going to over rule the computer and make the 17 go with the 24? They are both <300 rfu – They are both in the Danger Zone – No requirement for a phr to be >50% if Im in the Danger Zone

19. The Deconvolution So if I just lower the phr filter to 40%, they go together

20. The Deconvolution I can now deconvolute that locus and send the results to my new table In fact, we get 4 loci with only one option for the Unknown type that auto-graph So far: – V = 6% – C = 68% – U = 27%

21. The Deconvolution

22. D18 – This is really hard to see, so I apologize – 13, 15 is best (15, 15 P = 14%) (14, 15 phr 59%)

23. The Deconvolution D16 – A homozygote 9 (thats the only Unknown allele) means 11% Unknown rather than the 20+% weve been seeing

24. The Deconvolution D16

25. The Deconvolution D16 – Welcome to the Not Excluded option Even by the eyeball method, you can see that a 9, 11 works well for Unknown

26. The Deconvolution D16 (not excluded) option Circular or double sharing

27. The Egram

28. The Deconvolution D16 – Test the not excluded option by raising PHR to 70% Its the only thing left – best fit But since no P, it cant graph – (Maybe some day – weve got some ideas…)

29. The Deconvolution D5 V = 11, 13 C = 8, 12 (Major) U= ? (mid level)

30. The Deconvolution D5, U = ? (mid level) – But lots of choices

31. The Deconvolution D5 – mid-level has averaged 27% so far ?

32. The Deconvolution D5 set mP to 5% to get rid of the tiny bits Also set PHR to 60% to get rid of the option with the Unknown type being the 89% major contributor

33. The Deconvolution D5 now only 2 choices I could live with this using the obligate function, but…. Wheres the fun in that? At 70% PHR, the Unknown must be 12, 12 – That P of 26% is very nice compared to 27% avg – PHRs for the 2 heterozygous knowns are still OK

34. The Deconvolution D7 V = 7, 10 C = 8, 10 Pretty sure U has a 9

35. The Deconvolution D7 – 9 is in sutter, so correct for it Weve learned 50% correction is OK

36. The Deconvolution D7 – What did stutter correction do for us? Weve learned from testing that the true combination generally benefits the most from stutter correction The false combination may not show improvement It can be rather subtle (Im talking about PHR and P)

37. The Deconvolution D7 – Before correction 7,10(phr=98; p=07) 8,10(phr=98; p=79) 9(p=15) 7,10(phr=76; p=06) 8,10(phr=76; p=68) 9,10(phr 76; p=25) D7 – After correction 7,10(phr=93; p=04) 8,10(phr=93; p=83) (p=13) 7,10(phr=81; p=04) 8,10(phr=81; p=72) 9,10(phr 81; p=24)

38. The Deconvolution D7 – So the stutter correction generally improved the bottom option, and (slightly) unimproved the middle option At 80% phr, the top (not excluded) option went away (I admit the not excluded takes some practice samples to get comfortable with)

39. The Deconvolution D7 – My final answer Because the bottom answer improved the PHR fairly well and kept the P (24%) of the Unknown right where its been (27% avg), I pick 9, 10 – I know that both choices have good PHR – I know that I picked the one with the 2 nd best PHR – But, I cant figure out why the enzyme would miss half of the Unknown DNA (I didnt choose the one with P = 13%)

40. The Deconvolution Where we stand so far:

41. The Egram

42. The Deconvolution THO1 – V is 7, 9 E is 6, 8 7, 7 would work great for a mid-level

43. The Deconvolution In fact, at 70% PHR, the Unknown is a 7, 7 (25% of total DNA) Great PHR for the other two – 100% and 95%

44. The Deconvolution D13 Again, why would I pick an option where ½ the Unknown DNA just disappeared? This tells me hes not a homozygote here

45. The Deconvolution CSF – Kind of a mess to start with

46. The Deconvolution CSF – But V is the low level, not 39%

47. The Deconvolution CSF – Look carefully at these options The top one says Consensual and Unknown together combine to account for 61% - meaning V is 39% - doesnt make fit The bottom one says Victim and Unknown combine to account for 39%

48. The Deconvolution CSF – Quite a few choices, so I really have to study the peak pattern and the known profiles

49. The Deconvolution CSF – This tells me that the Victim is totally masked

50. The Deconvolution CSF In fact, Vs 6% is probably lost in the expected variation of the other 2 contributors Therefore, I will temporarily un-apply the Victim as a required genotype, and treat this as a 2 person locus

51. The Deconvolution CSF As a 2 person locus, 11, 12 isnt bad

52. The Deconvolution CSF Then correcting for 50% stutter pretty much makes it perfect

53. The Deconvolution CSF So Ill go back to a three person mixture and go with an 11, 12

54. The Egram

55. The Deconvolution D8 V = 14, 14 C = 13, 14

56. The Deconvolution D8 – Change to two person locus as before, Victim + Unknown around 30% so far So could Unknown be a 14, 14 (only 14%)? But the 14 Allele is 8642 rfu (blown out)

57. The Deconvolution D8 – Since 14 allele saturated the detector, we dont know what the real height is A bunch of 14 could still be out there somewhere So even though a 14, 14 looks like a low proportion, thats my call (You should probably have an upper limit!) In our current world Id have to really argue to use this injection – this is an old, old sample

58. The Deconvolution TPOX V = 8, 8 C = 8, 11

59. The Deconvolution TPOX 1% for V + U – NO 99% for C + U – Maybe, but V is pretty good at D16, D18 and D2 so probably not

60. The Deconvolution TPOX We can test by raising PHR Only 11, 11 remains

61. The Deconvolution D3 V = 15, 16 E = 15, 16 Finally an easy one 15, 15 (12%) and 16, 16 (1%) dont work – 15, 16

62. The Deconvolution That leaves FGA We know V drops out here So – In the following order: – Un-apply V reference (we know the 22 dropped) – Ignore the 20 allele (it has nothing to do with the other two alleles) – Switch to 2 person calculator – (You can always consider an allele again)

63. The Deconvolution FGA Might seem easy – its not 1:1 But, this is worst case scenario for stutter! 24 is very tall, and stutter at FGA = 17%

64. The Deconvolution FGA So correct for stutter at FGA Use the 50% we know fits mixtures 23, 24 for Unknown is 34%, Consensual is 66%

65. The Deconvolution We now have a full single source Unknown profile – no Anys, no Obligates

66. The Deconvolution Graph looks good, but some things dont graph D8S1179 CSF1PO D3S1358 Amel FGA 2 people share type 3 people share typeNot Excluded 2 people share typeVictim dropped out

67. The Stat We cant really use the option from the stat window cause the software remembers a mess – Some loci were converted to 2 contributors – We ignored an allele at FGA – Etc…

68. The Stat But, who cares? We have a single source profile 1 in 124 Quintillion

69. How many of you would… Countdown 30 0 of 30 1.Say I was nuts to do that deconvolution? 2.Agree and be OK as my Tech Reviewer? 3.Want to go talk to my Tech Leader about my interpretation?

70. How many think… Countdown 30 0 of 30 1.I was conservative? 2.I was the opposite of conservative? 3.I got the right answer?

71. Match to Suspect I call that an exclusion Is it still an anti-conservative interpretation if I excluded the guy?

72. Now What? Well, because we excluded this suspect, they sent in another reference from some other guy that was at the party This first guy always said that he went in her room, but it was only to get his shoes that he left in there So now well bring in Suspect #2s reference and see how it fits

73. New Data Table To cut down on open windows, well do a new table of the original mixture, the Unknown type and the new reference of Suspect #2

74. Match to Suspect #2 Suspect #2 matches the Unknown profile we deconvoluted from the mixture

75. A Much Faster Way Apply all three references, V, C, and S Set for 3 people Auto call references View call report

76. A Much Faster Way Doesnt graph well – Click through the loci There are 7 with zero support (no combinations)

77. A Much Faster Way Get on the phone to the investigators and tell them that although all of Suspects DNA is found in the mixture, it just doesnt fit right. Can you please send in another reference? Then apply that new reference to the mixture and see how it fits

78. A Much Faster Way Still doesnt graph every where, but we know that (shared types, not excluded, etc) But support at every locus (D2 and FGA special)

79. Things to Consider It takes more than just the presence of alleles to be an inclusion The inclusion should be supported by all PHR and P expectations – its about the math I know were supposed to look at the mixture without applying the reference, but…

80. Things to Consider If you apply all references and nothing fits, you have an exclusion If you apply the references and it all fits, you just did a real time LR (sort of) But most importantly:

81. Things to Consider Even though we peeked at the reference of Suspect #1, it played no role whatsoever in the final interpretation and stat If we would have stopped when we saw the result of the first comparison of Suspect #1 to the mixture and just done a CPI, we would have just included an innocent man