Figure 1.
Cell suspensions created in various suspension mediums. 1× PBS (red), nuclease-free water (blue), and TE−4 (green). 1C = 1 cell, 2C = 2 cells, n = number of samples analyzed.
Figure 1.
Cell suspensions created in various suspension mediums. 1× PBS (red), nuclease-free water (blue), and TE−4 (green). 1C = 1 cell, 2C = 2 cells, n = number of samples analyzed.
Figure 2.
DSCS D16S539 back stutter.
Figure 2.
DSCS D16S539 back stutter.
Figure 3.
Applied BiosystemsTM 3500 Genetic Analyzer saturation threshold (DSCS) (A) Standard Analysis (B) DSCS. Compared to standard analysis, DSCS samples have a greater discrepancy in observed and expected allele heights due to elevated stutter. Therefore, instrument saturation limits should be determined using standard analysis.
Figure 3.
Applied BiosystemsTM 3500 Genetic Analyzer saturation threshold (DSCS) (A) Standard Analysis (B) DSCS. Compared to standard analysis, DSCS samples have a greater discrepancy in observed and expected allele heights due to elevated stutter. Therefore, instrument saturation limits should be determined using standard analysis.
Figure 4.
Extreme peak height imbalance possible with single cell analysis illustrated with the locus D22S1045. The known genotype of the cell donor is 11, 16 (10 = back stutter).
Figure 4.
Extreme peak height imbalance possible with single cell analysis illustrated with the locus D22S1045. The known genotype of the cell donor is 11, 16 (10 = back stutter).
Figure 5.
DSCS peak height variability (A) Standard Analysis (B) DSCS. DSCS results in a broader distribution due to the increased variance/peak height imbalance due to low template effects.
Figure 5.
DSCS peak height variability (A) Standard Analysis (B) DSCS. DSCS results in a broader distribution due to the increased variance/peak height imbalance due to low template effects.
Figure 6.
DSCS post burn-in accepts.
Figure 6.
DSCS post burn-in accepts.
Figure 7.
DSCS (3 cell subsample) inter-locus peak height variance.
Figure 7.
DSCS (3 cell subsample) inter-locus peak height variance.
Figure 8.
DSCS (3 cell subsample) decrease in locus specific amplification efficiency at inhibited loci (boxed and asterisked).
Figure 8.
DSCS (3 cell subsample) decrease in locus specific amplification efficiency at inhibited loci (boxed and asterisked).
Figure 9.
DSCS (3 cell subsample, S3-5C-1) degradation curves. Regular sample (A), Degraded sample (B), Inhibited sample (C).
Figure 9.
DSCS (3 cell subsample, S3-5C-1) degradation curves. Regular sample (A), Degraded sample (B), Inhibited sample (C).
Figure 10.
Sensitivity and specificity of DSCS (EFM) analysis with single source subsamples. LR = 0 plotted as −350. Green circles (1-cell known contributor), yellow circles (2-cell known contributor), orange circles (non-contributor). Total results (A); False positives (i.e., log (LR) > 0) (B).
Figure 10.
Sensitivity and specificity of DSCS (EFM) analysis with single source subsamples. LR = 0 plotted as −350. Green circles (1-cell known contributor), yellow circles (2-cell known contributor), orange circles (non-contributor). Total results (A); False positives (i.e., log (LR) > 0) (B).
Figure 11.
Specificity of DSCS (EFM) mini-mixture analysis. EuroForMix; Single source subsamples. LR = 0 plotted as −50. Green circles (known contributors), orange circles (non-contributor).
Figure 11.
Specificity of DSCS (EFM) mini-mixture analysis. EuroForMix; Single source subsamples. LR = 0 plotted as −50. Green circles (known contributors), orange circles (non-contributor).
Figure 12.
Overestimating the number of contributors to single source DSCS subsamples. STRmixTM (A); EuroForMix (B); Note: Non-contributor profiles were not run with EFM. LR = 0 plotted as −50.
Figure 12.
Overestimating the number of contributors to single source DSCS subsamples. STRmixTM (A); EuroForMix (B); Note: Non-contributor profiles were not run with EFM. LR = 0 plotted as −50.
Figure 13.
Drop-out from a single heterozygous TPOX allele. ADO = allele drop-out; * = pull-up. Stutter was retained for the sample when analyzed with PG.
Figure 13.
Drop-out from a single heterozygous TPOX allele. ADO = allele drop-out; * = pull-up. Stutter was retained for the sample when analyzed with PG.
Figure 14.
Underestimating the number of contributors to 2-cell DSCS mini-mixtures. (A) STRmixTM; (B) EuroForMix; Note: Non-contributor profiles were not run with EFM. LR = 0 plotted as −50.
Figure 14.
Underestimating the number of contributors to 2-cell DSCS mini-mixtures. (A) STRmixTM; (B) EuroForMix; Note: Non-contributor profiles were not run with EFM. LR = 0 plotted as −50.
Figure 15.
Specificity improvement of DSCS (EFM) with replicate analysis. Known contributors above the diagonal (i.e., y = x) represent improved LR recovery from replicate analysis. Single Source Cell Replicates. LR = 0 plotted as −350. Green circles (known contributors); orange circles (non-contributors). (A) All samples. (B) Detail showing log(LR) > 0 samples.
Figure 15.
Specificity improvement of DSCS (EFM) with replicate analysis. Known contributors above the diagonal (i.e., y = x) represent improved LR recovery from replicate analysis. Single Source Cell Replicates. LR = 0 plotted as −350. Green circles (known contributors); orange circles (non-contributors). (A) All samples. (B) Detail showing log(LR) > 0 samples.
Figure 16.
Specificity of DSCS (EFM) mini-mixture analysis. LR = 0 plotted as −350. Green circles (known contributors), orange circles (non-contributor) (A) All samples. (B) Detail showing log(LR) > 0 samples.
Figure 16.
Specificity of DSCS (EFM) mini-mixture analysis. LR = 0 plotted as −350. Green circles (known contributors), orange circles (non-contributor) (A) All samples. (B) Detail showing log(LR) > 0 samples.
Figure 17.
Increased contributor log(LR) recovery in 2–4 person mixtures by DSCS (EFM) (“DSCS Replicate”) compared to standard PG mixture analysis (STD Mix). The “Reference” LR is the reciprocal of the random match probability of the single source DNA profile from the known donor. The alphanumeric characters refer to the individual donors present in the mixture.
Figure 17.
Increased contributor log(LR) recovery in 2–4 person mixtures by DSCS (EFM) (“DSCS Replicate”) compared to standard PG mixture analysis (STD Mix). The “Reference” LR is the reciprocal of the random match probability of the single source DNA profile from the known donor. The alphanumeric characters refer to the individual donors present in the mixture.
Figure 18.
Increased contributor log(LR) recovery in 5 (left panel) and 6 (right panel) person mixtures by DSCS (EFM) (“DSCS Replicate”). Standard PG mixture analysis (“STD Mix”) could not be conducted on these mixtures due to EFM software limitations. The “Reference” LR is the reciprocal of the random match probability of the single source DNA profile from the known donor. The alphanumeric characters refer to the individual donors present in the mixture.
Figure 18.
Increased contributor log(LR) recovery in 5 (left panel) and 6 (right panel) person mixtures by DSCS (EFM) (“DSCS Replicate”). Standard PG mixture analysis (“STD Mix”) could not be conducted on these mixtures due to EFM software limitations. The “Reference” LR is the reciprocal of the random match probability of the single source DNA profile from the known donor. The alphanumeric characters refer to the individual donors present in the mixture.
Figure 19.
Contributor log (LR) recovery by DSCS (EFM) (“DSCS Replicate”) in an additional 6-person mixture. The “Reference” LR is the reciprocal of the random match probability of the single source DNA profile from the known donor. Standard PG mixture analysis (“STD Mix”) could not be conducted on the mixture due to EFM software limitations. The alphanumeric characters refer to the individual donors present in the mixture.
Figure 19.
Contributor log (LR) recovery by DSCS (EFM) (“DSCS Replicate”) in an additional 6-person mixture. The “Reference” LR is the reciprocal of the random match probability of the single source DNA profile from the known donor. Standard PG mixture analysis (“STD Mix”) could not be conducted on the mixture due to EFM software limitations. The alphanumeric characters refer to the individual donors present in the mixture.
Table 1.
Allele recovery of 1- and 2-cell subsamples collected after 6 months from either a cell suspension or a saliva stain.
Table 1.
Allele recovery of 1- and 2-cell subsamples collected after 6 months from either a cell suspension or a saliva stain.
| 1-Cell Subsamples | 2-Cell Subsample |
---|
Saliva Stain | 25 ± 12 | 29 ± 10 |
Cell Suspension | 22 ± 12 | 26 ± 14 |
Table 2.
STRmixTM drop-in data/parameters.
Table 2.
STRmixTM drop-in data/parameters.
Distribution | Drop-In Events | Samples Analyzed | Loci | Samples × Loci | Drop-In Rate | Drop-In Cap |
---|
Uniform | 11 | 35 | 21 | 735 | 0.0164 | 30,000 RFU |
Table 3.
DSCS parameters test.
Table 3.
DSCS parameters test.
Sample | EFM Log(LR) |
---|
S5-3C-1 | 27.55 |
S5-3C-1 drop-in | 27.55 |
S5-3C-1 inhibited | 27.55 |
S5-3C-1 degraded | 27.43 |
Table 4.
Percentage of subsamples and replicates analyzed with EuroForMix that reach the specified threshold.
Table 4.
Percentage of subsamples and replicates analyzed with EuroForMix that reach the specified threshold.
Sample Type | Subsample | Replicate | Subsample | Replicate |
---|
log(LR) > 0 | log(LR) ≥ 6 |
---|
SS 1 cell (s = 100) | 87% | 100% | 45% | 94% |
SS 2 cell (s = 81) | 89% | 100% | 64% | 93% |
Mix 2 cell (s = 110) | 60% | 100% | 30% | 93% |
Table 5.
Misclassification of single source subsamples for replicate analysis.
Table 5.
Misclassification of single source subsamples for replicate analysis.
Correct Profile S3 | Incorrect Profile Included in Replicate (Various Donors) |
---|
Sample | Total Alleles | STRmixTM Replicate log(LR) | EFM Replicate log(LR) | Misclassified Sample | Total Alleles | Alleles That Do Not Match S3 | STRmixTM Replicate log(LR) S3 | EFM Replicate log(LR) S3 |
---|
S3-6 | 19 | 14 | 12 | CM31 | 4 | 2 | 14 | 12 |
S3-9 | 18 | SA10 | 7 | 5 | 0 | −20 |
| | | | S5-24 | 14 | 6 | −9 | −19 |
| | | | S5-25 | 21 | 11 | 0 | −3 |
| | | | S8 | 32 | 19 | 0 | −101 |
Table 6.
Misclassification of mini-mixtures for replicate analysis.
Table 6.
Misclassification of mini-mixtures for replicate analysis.
Correct Profiles S5/CM31 Mini-Mixture | Incorrect Profile Included in Replicate (Various Donors) |
---|
Sample | STRmix S5 | STRmix CM31 | EFM S5 | EFM CM31 | Misclassified Sample | Total Alleles | Alleles That Do Not Match S5 | Alleles That Do Not Match CM31 | STRmix Replicate log(LR) S5 | STRmix Replicate log(LR) CM31 | EFM Replicate log(LR) S5 | EFM Replicate log(LR) |
---|
S5CM31-4 | 9 | 16 | 2 | 14 | SA10 | 7 | 5 | 3 | 9 | 16 | 8 | FAIL * |
S5CM31-6 | 1 | 15 | 3 | 16 | S3 | 19 | 11 | 13 | 0 | 16 | −4 | 15 |
S5CM31-28 | 8 | 10 | 9 | 11 | S8 | 32 | 20 | 18 | 0 | 16 | −30 | FAIL * |
Replicate | 10 | 17 | 8 | 18 | S3/SA10-17 | | | | 0 | 10 | −12 | 15 |
S3/SA10-25 | | | | 4 | 16 | FAIL * | FAIL * |
S3/SA10-17&25 | | | | 0 | 5 | FAIL * | FAIL * |