NIPT Technique Based on the Use of Long Chimeric DNA Reads

• Published in: Genes Vol. 11; no. 6; p. 590
• Main Authors: Belova, Vera, Plakhina, Daria, Evfratov, Sergey, Tsukanov, Kirill, Khvorykh, Gennady, Rakitko, Alexander, Konoplyannikov, Alexander, Ilinsky, Valery, Rebrikov, Denis, Korostin, Dmitriy
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI 26.05.2020; MDPI AG
• Subjects: Adult; Aneuploidy; Cell-Free Nucleic Acids - blood; Cell-Free Nucleic Acids - genetics; cfDNA; Chimera - genetics; chimeric DNA; Chromosomes, Human, Pair 13 - genetics; Chromosomes, Human, Pair 18 - genetics; Chromosomes, Human, Pair 21 - genetics; Female; fetal fraction; Humans; NGS; NIPT; Pregnancy; Prenatal Diagnosis; short DNA fragments; Trisomy 13 Syndrome - blood; Trisomy 13 Syndrome - genetics; Trisomy 13 Syndrome - pathology; Trisomy 18 Syndrome - blood; Trisomy 18 Syndrome - genetics; Trisomy 18 Syndrome - pathology; short DNA fragments; long chimeric reads; cfDNA; NGS; NIPT; chimeric DNA; fetal fraction
• ISSN: 2073-4425; 2073-4425
• DOI: 10.3390/genes11060590

Non-invasive prenatal testing (NIPT) for aneuploidy on Chromosomes 21 (T21), 18 (T18) and 13 (T13) is actively used in clinical practice around the world. One of the limitations of the wider implementation of this test is the high cost of the analysis itself, as high-throughput sequencing is still relatively expensive. At the same time, there is an increasing trend in the length of reads yielded by sequencers. Since extracellular DNA is short, in the order of 140-160 bp, it is not possible to effectively use long reads. The authors used high-performance sequencing of cell-free DNA (cfDNA) libraries that went through additional stages of enzymatic fragmentation and random ligation of the resulting products to create long chimeric reads. The authors used a controlled set of samples to analyze a set of cfDNA samples from pregnant women with a high risk of fetus aneuploidy according to the results of the first trimester screening and confirmed by invasive karyotyping of the fetus using laboratory and analytical approaches developed by the authors. They evaluated the sensitivity, specificity, PPV (positive predictive value), and NPV (negative predictive value) of the results. The authors developed a technique for constructing long chimeric reads from short cfDNA fragments and validated the test using a control set of extracellular DNA samples obtained from pregnant women. The obtained sensitivity and specificity parameters of the NIPT developed by the authors corresponded to the approaches proposed earlier (99.93% and 99.14% for T21; 100% and 98.34% for T18; 100% and 99.17% for T13, respectively).