Peak Height Ratios in Peak Height Ratios in Forensic STR Analyses:Forensic STR Analyses:

Pattern of Occurrence & Effects of Pattern of Occurrence & Effects of ConcentrationConcentration

Data from:Data from: Illinois State Police Laboratory Illinois State Police Laboratory Broward County Sheriff’s Lab & Broward County Sheriff’s Lab &

Forensic BioinformaticsForensic Bioinformatics

69%

ValidationValidation

Size Ladders are Used to Make Allele Calls

Locus N Mean%ABI

SD 99%CI N Mean%FBI

SD 99%CI

D3S1358 68 93 4 81 167193

9389

5.29.4

7762

VWA 74 93 5 78 180 92 5.6 75

D16S539 70 92 6 74 220 88 9.4 60

FGA 80 93 5 78 269 92 5.7 75

THO1 70 92 6 74 318 91 8.0 67

TPOX 78 92 6 74 298 88 8.6 63

CSF1PO 84 92 6 74 234 88 9.1 61

D8S1179 93 92 6 74 180 93 4.8 79

D21S11 95 91 7 70 230 92 5.5 76

D18S51 100 91 6 73 271 89 6.7 69

D5S818 65 92 5 77 122 93 4.9 78

D13S317 73 93 5 78 173 93 5.6 76

D7S820 79 93 6 75 159261

9288

5.99.4

7460

Figure from: STR ANALYSIS BY CAPILLARY ELECTROPHORESIS: DEVELOPMENT OF INTERPRETATION GUIDELINES FOR THE PROFILER PLUS™ AND COFILER™ SYSTEMSFOR USE IN FORENSIC SCIENCE by Deborah Hobson, Jill Smerick, and Jenifer Smith DNAU I, FBI Laboratory, Washington, D.C.

Scatterplot (Spreadsheet in phril.stw 9v*962c)

-1000 0 1000 2000 3000 4000 5000 6000 7000 8000

RFU2

40

50

60

70

80

90

100

110

PH

R

RFU values for smaller allele in each heterozygote

Frequency table: PHR: =(V4/V3)*100 (PHRATIO.STA in phril.stw)K-S d=.10275, p<.01 ; Lilliefors p<.01

CategoryCountCumulative

CountPercentof Valid

Cumul %of Valid

% of allCases

Cumulative %of All

40.00000<x<=50.0000050.00000<x<=60.0000060.00000<x<=70.0000070.00000<x<=80.0000080.00000<x<=90.0000090.00000<x<=100.0000Missing

2 2 0.23838 0.23840.23838 0.238410 12 1.19190 1.43031.19190 1.430327 39 3.21812 4.64843.21812 4.648479 118 9.4159714.06449.41597 14.0644

275 39332.7771246.841532.77712 46.8415446 83953.15852100.000053.15852 100.0000

0 839 0.00000 0.00000 100.0000

Frequency of Occurrence of Heterozygote Peak Height Ratios in Percentages in All Loci in the Illinois State Police Validation Studies

Histogram: PHR: =(V4/V3)*100K-S d=.10275, p<.01 ; Lilliefors p<.01

Expected Normal

40 50 60 70 80 90 100

X <= Category Boundary

0

50

100

150

200

250

300

350

400

450

500

No. of obs.

Plot of Means and Conf. Intervals (95.00%)Validation Samples

D3 vWA FGA AMEL D8 D21 D18 D5 D13 D7 D16 THO1 TPOX CSF

LOCUS

78

80

82

84

86

88

90

92

94

96

Values in P

ercent

Mean PHR with 95% CI for Each CODIS Locus

D7 CSF D3 D16 TPOX D8 D21 D13 vWA D18 D5 FGA THO1

Locus

0.86

0.87

0.88

0.89

0.90

0.91

0.92

0.93

PH

R

Reference Sample Peak Height Ratios

Median & Range PHR/Locus

D7 CSF D3 D16 TPOX D8 D21 D13 vWA D18 D5 FGA THO1

Locus

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

1.1

PH

R

2-Way Tables of Descriptive Statistics (PHRATIO.STA in phril.stw)N=839 (No missing data in dep. var. list)

RFUClassPHR

MeansPHRN

PHRStd.Dev.

PHRMinimum

PHRMaximum

< 500rfu500<rfu<10001000-2000rfu2000-4000rfu> 4000rfuAll Grps

85.0742516711.6051146.80233100.000089.30670164 7.4422662.65244 99.627689.56659189 8.6625457.27371 99.843889.94837164 7.1586858.04441 99.907489.40954155 8.2854857.15089 99.906288.66722839 8.9514646.80233100.0000

Analysis of Variance (PHRATIO.STA in phril.stw)Marked effects are significant at p < .05000

VariableSS

Effectdf

EffectMS

EffectSS

Errordf

ErrorMSError

F p

PHR 2730.409 4 682.602464417.4183477.239108.8375230.000001

Scheffe Test; Variable: PHR (PHRATIO.STA in phril.stw)Marked differences are significant at p < .05000

RFUClass{1}

M=85.074{2}

M=89.307{3}

M=89.567{4}

M=89.948{5}

M=89.410< 500rfu {1}500<rfu<1000 {2}1000-2000rfu {3}2000-4000rfu {4}> 4000rfu {5}

0.0007880.0001340.0000480.0006680.000788 0.9992800.9792900.9999850.0001340.999280 0.9967450.9999080.0000480.9792900.996745 0.9898240.0006680.9999850.9999080.989824

The mean Peak Height Ratio is significantly smaller and the variance larger for smaller DNA samples as reflected by the mean RFU of a sample. Above that threshold the mean PHR & Variance does not differ significantly among peak height classes.

Relationship Between DNA Amount and Allele Imbalance

< 500rfu500<rfu<1000

1000-2000rfu2000-4000rfu

> 4000rfu

Mean RFU per Sample

83

84

85

86

87

88

89

90

91

92

Mea

n P

eak

Hei

ght R

atio

Mean ±SE ±1.96*SE

Summary Frequency Table (PHRATIO.STA in phril.stw)Marked cells have counts > 10(Marginal summaries are not marked)

RFUClass 70%Yes

70%No

RowTotals

CountColumn PercentRow PercentTotal PercentCountColumn PercentRow PercentTotal PercentCountColumn PercentRow PercentTotal PercentCountColumn PercentRow PercentTotal PercentCountColumn PercentRow PercentTotal PercentCountTotal Percent

< 500rfu 150 17 16718.63%50.00%89.82%10.18%17.88%2.03%19.90%

500<rfu<1000 162 2 16420.12%5.88%98.78%1.22%19.31%0.24%19.55%

1000-2000rfu 184 5 18922.86%14.71%97.35%2.65%21.93%0.60%22.53%

2000-4000rfu 160 4 16419.88%11.76%97.56%2.44%19.07%0.48%19.55%

> 4000rfu 149 6 15518.51%17.65%96.13%3.87%17.76%0.72%18.47%

All Grps 805 34 83995.95%4.05%

Relationship between RFU & thepercent of profiles with PHR’sabove (Yes) and below (NO) 70%.

Chi-Squared = 21.58, 4df, p = 0.00024

Percent of Samples with PHR Less Than 70% as a Function of the Mean RFU of the Sample in the Illinois State Police Crime Lab Validation

Studies

0

2

4

6

8

10

12

<500 500<Y<1000 1-2000 2-4000 >4000

Mean RFU Values per Sample

Perc

ent o

f PH

R <

70%

Peak Height Ratios for Validation Samples

0 1 2 3 4 5 6 7 8 9

Concentration Class

66

68

70

72

74

76

78

80

82

84

86

88

90

92

94

PH

R as P

ercent

Mean

Mean±SE

Mean±1.96*SE

Reference Samples

0 1 2 3 4 5 6 7

Concentration Class

0.86

0.87

0.88

0.89

0.90

0.91

0.92

0.93

PH

R as P

ercent

Mean Mean±SE Mean±1.96*SE

Amplifications from same dilution tube (~30 pg). (2-012802 10.19 AM)

Allelic imbalance present. Allele dropout @ D18.“Different DNA profiles” @ D3, D21, D18 and D13.

The bottom line is simple; Peak Height Ratios (PHR’s) have a higher probability of being imbalanced and show significantly greater imbalances in small and/or degraded samples of DNA when using Profiler Plus and Cofiler to type STR DNA in crime scene type samples. A significant number of samples at all RFU classes show peak height ratios below 70%. This is especially true of small and/or degraded samples like those collected at many crime scenes. The Illinois State Police DNA lab’s use of a 70% rule to infer single source samples and in interpreting the relative contributions to mixtures is not justified by the data in their own “validation” studies. I have seen similar data from the validation studies of many other crime labs (e.g., Minnesota BCA, Broward County Sheriffs Lab, Bode Technology, West Palm Beach Sheriffs Office, Connecticut State Police CrimeLab, FBI Laboratory, and studies done by the kit’s manufacturers.