Fig. 5: The use of cfDNA characteristics for the detection of fetal monogenic variants. | Cell Discovery

Fig. 5: The use of cfDNA characteristics for the detection of fetal monogenic variants.

From: Genetic deconvolution of fetal and maternal cell-free DNA in maternal plasma enables next-generation non-invasive prenatal screening

Fig. 5

a The identification of fetal monogenic variants includes ROI analysis, ACD filtering, and FMID filtering. b Fetal-Maternal Nearest Neighbor Insert-size Calibration was first used to exclude those reads harboring wild-type alleles which possess the closest cfDNA fragment length to the reads harboring the variant alleles of a potential fetal origin. The remaining fragments with the wild-type allele were compared with those with the variant alleles for their lengths to identify potential fetal SNVs. c Different insert-size distribution for wild-type (ref) and variant (alt) allele supporting reads on 28 samples tested. For all variants detected, median insert-size of ref (RefinsMid) and alt (AltinsMid) allele supporting reads were box plotted with upper whisker (Q3 + 1.5 × IQR), Q3, Q2, average, Q1, and lower whisker (Q1 – 1.5 × IQR) to demonstrate the differences between the TP or FP variants. Insert-size was ~10 bp shorter (P < 1.0 × 10−15, two-tailed unequal-variance t-test) in alt allele group compared to ref allele group on TP variants consistent with a fetal origin and no such difference was seen in FP variants. d, e Sensitivity (d) and PPV (e) comparison for different filtering methods using 28 validation samples. By applying both the ACD and FMID variant filters, the test sensitivity was essentially unchanged at 99.5% while the PPV was significantly improved (P < 0.01). When only the ACD filter was used, the test sensitivity was reduced to 96.8% (P < 1.0 × 10−8). Upper whisker (Q3 + 1.5 × IQR), Q3, Q2, average, Q1, lower whisker (Q1 – 1.5 × IQR), and all non-outlier data points between lower and upper whiskers were demonstrated on the box plot. The ACD and FMID filters were used to filter in variants of a likely fetal origin. Ref reference, Alt alternative, TP true positive, FP false positive, Q1 lower quartile, Q2 median quartile, Q3 upper quartile, IQR inter-quartile range = Q3 – Q1; ROI regions of interest, ACD allele count distribution, FMID fetal-maternal insert-size distribution, PPV positive predictive value, FF fetal fraction, AF allelic fraction Min P-value, minimum of the four P-values to examine whether alternative allele fragments are significantly different from the reference allele fragments in length. CDF, the absolute value of the log cumulative distribution function value; NoFLT no variant filter applied.

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