Abstract
Concerns of traditional prenatal aneuploidy testing methods, such as low accuracy of noninvasive and health risks associated with invasive procedures, were overcome with the introduction of novel noninvasive methods based on genetics (NIPT). These were rapidly adopted into clinical practice in many countries after a series of successful trials of various independent submethods. Here we present results of own NIPT trial carried out in Moscow, Russia. 1012 samples were subjected to the method aimed at measuring chromosome coverage by massive parallel sequencing. Two alternative approaches are ascertained: one based on maternal/fetal differential methylation and another based on allelic difference. While the former failed to provide stable results, the latter was found to be promising and worthy of conducting a large-scale trial. One critical point in any NIPT approach is the determination of fetal cell-free DNA fraction, which dictates the reliability of obtained results for a given sample. We show that two different chromosome Y representation measures-by real-time PCR and by whole-genome massive parallel sequencing-are practically interchangeable (r=0.94). We also propose a novel method based on maternal/fetal allelic difference which is applicable in pregnancies with fetuses of either sex. Even in its pilot form it correlates well with chromosome Y coverage estimates (r=0.74) and can be further improved by increasing the number of polymorphisms.
Highlights
Aneuploidies can be attributed to cause 30% of miscarriage cases, and affect up to 1 in 300 live births[1]
A new noninvasive prenatal testing technology based on sequencing of cell-free DNA from maternal blood was widely implemented in the industry
Trisomy of a certain chromosome in the fetus may be detected through sequencing of total cfDNA from maternal blood plasma and subsequent counting of reads mapped on each chromosome
Summary
Aneuploidies can be attributed to cause 30% of miscarriage cases, and affect up to 1 in 300 live births[1]. Blood plasma of a pregnant woman contains cell-free DNA fragments of both maternal and fetal origin. The latter permeates through the placental barrier into the main blood flow. Fetal fraction makes up 10–20% of all blood plasma cfDNA on average, rising through the whole pregnancy duration. Trisomy of a certain chromosome in the fetus may be detected through sequencing of total cfDNA from maternal blood plasma and subsequent counting of reads mapped on each chromosome. Such chromosome would show statistically significant increase in coverage[5,6,7]. Underrepresentation of fetal genetic material might lead to false negative outcomes, so such cases must be diagnosed by other means
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