Abstract
Massively parallel sequencing (MPS) combined with bioinformatic analysis has been widely applied to detect fetal chromosomal aneuploidies such as trisomy 21, 18, 13 and sex chromosome aneuploidies (SCAs) by sequencing cell-free fetal DNA (cffDNA) from maternal plasma, so-called non-invasive prenatal testing (NIPT). However, many technical challenges, such as dependency on correct fetal sex prediction, large variations of chromosome Y measurement and high sensitivity to random reads mapping, may result in higher false negative rate (FNR) and false positive rate (FPR) in fetal sex prediction as well as in SCAs detection. Here, we developed an optimized method to improve the accuracy of the current method by filtering out randomly mapped reads in six specific regions of the Y chromosome. The method reduces the FNR and FPR of fetal sex prediction from nearly 1% to 0.01% and 0.06%, respectively and works robustly under conditions of low fetal DNA concentration (1%) in testing and simulation of 92 samples. The optimized method was further confirmed by large scale testing (1590 samples), suggesting that it is reliable and robust enough for clinical testing.
Highlights
Since cell-free fetal DNA was detected in cell-free DNA of pregnant women plasma by Lo et al in 1997 [1], comparison of chromosome dosage distribution of cffDNA between patient and control plays an increasingly important role in fetal aneuploidy diagnosis
These results suggest that these reads may not come from the Y chromosome but were randomly mapped in the female, because 1) DNA fragments from the Y chromosome should not be observed in sequencing data from the female; 2) the abundant variations and repeat DNA sequences in the Y chromosome could result in random mapping for short sequencing reads
We filtering out randomly mapping reads at the six special regions in the Y chromosome to achieve significantly smaller false negative rate (FNR) and false positive rate (FPR) of fetal sex detection than the current method [5, 6]
Summary
Since cell-free fetal DNA (cffDNA) was detected in cell-free DNA (cfDNA) of pregnant women plasma by Lo et al in 1997 [1], comparison of chromosome dosage distribution of cffDNA between patient and control plays an increasingly important role in fetal aneuploidy diagnosis. The non-invasive prenatal testing (NIPT) is based on shotgun massively parallel sequencing technology and has been widely applied to detect trisomy 21, 18, 13 and sex chromosome aneuploidies (SCAs). SCAs are characterized by an abnormal number of sex chromosomes and has been linked to a series of diseases, such as Turner syndrome (monosomy X, 1/ 2000 in baby), Klinefelter syndrome (XXY, 1/500 in male), Triple X syndrome Accurate Fetus Gender Prediction and Sex Chromosome Aneuploidy Detection funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript
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