Joaquim S L Vong1,2, Jason C H Tsang1,2, Peiyong Jiang1,2, Wing-Shan Lee1,2, Tak Yeung Leung3, K C Allen Chan1,2, Rossa W K Chiu1,2, Y M Dennis Lo4,2. 1. Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China. 2. Departments of Chemical Pathology and. 3. Obstetrics and Gynaecology, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China. 4. Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong SAR, China; loym@cuhk.edu.hk.
Abstract
BACKGROUND: Recent studies have suggested that single-stranded DNA (ssDNA) library preparation can enrich short DNA species from the plasma of healthy individuals, cancer patients, and transplant recipients. Based on previous observations that fetal DNA molecules in the maternal plasma are shorter than maternal DNA molecules, ssDNA library preparation may potentially enrich fetal DNA and provide substantial improvement in noninvasive prenatal testing. METHODS: We tested this hypothesis by comparing the maternal plasma DNA sequencing results using 2 types of ssDNA library preparation methods and a standard double-stranded DNA (dsDNA) library method using samples from first- and third-trimester pregnancies. We also evaluated the performance of ssDNA and dsDNA library methods in the noninvasive prenatal detection of trisomy 21 from maternal plasma. RESULTS: Short DNA species were significantly enriched in ssDNA libraries. However, contrary to previous speculation, no significant enrichment was observed in the overall fetal fraction in maternal plasma collected in the first trimester. Our use of an ssDNA library did not reduce the variation in chromosomal representation when compared with a standard dsDNA library in the first-trimester plasma samples. ssDNA libraries also showed inferior performance in the noninvasive prenatal detection of trisomy 21 from maternal plasma. Detailed fetal fraction analysis using size-fractionated Y chromosome sequences and fetal-specific single-nucleotide polymorphisms (SNPs) revealed an unexpected finding that short maternal DNA was preferentially enriched over short fetal DNA in an ssDNA library irrespective of GC content. CONCLUSIONS: Our findings have shown that ssDNA library preparation preferentially enriches short maternally derived DNA in maternal plasma.
BACKGROUND: Recent studies have suggested that single-stranded DNA (ssDNA) library preparation can enrich short DNA species from the plasma of healthy individuals, cancerpatients, and transplant recipients. Based on previous observations that fetal DNA molecules in the maternal plasma are shorter than maternal DNA molecules, ssDNA library preparation may potentially enrich fetal DNA and provide substantial improvement in noninvasive prenatal testing. METHODS: We tested this hypothesis by comparing the maternal plasma DNA sequencing results using 2 types of ssDNA library preparation methods and a standard double-stranded DNA (dsDNA) library method using samples from first- and third-trimester pregnancies. We also evaluated the performance of ssDNA and dsDNA library methods in the noninvasive prenatal detection of trisomy 21 from maternal plasma. RESULTS: Short DNA species were significantly enriched in ssDNA libraries. However, contrary to previous speculation, no significant enrichment was observed in the overall fetal fraction in maternal plasma collected in the first trimester. Our use of an ssDNA library did not reduce the variation in chromosomal representation when compared with a standard dsDNA library in the first-trimester plasma samples. ssDNA libraries also showed inferior performance in the noninvasive prenatal detection of trisomy 21 from maternal plasma. Detailed fetal fraction analysis using size-fractionated Y chromosome sequences and fetal-specific single-nucleotide polymorphisms (SNPs) revealed an unexpected finding that short maternal DNA was preferentially enriched over short fetal DNA in an ssDNA library irrespective of GC content. CONCLUSIONS: Our findings have shown that ssDNA library preparation preferentially enriches short maternally derived DNA in maternal plasma.
Authors: Irena Hudecova; Christopher G Smith; Robert Hänsel-Hertsch; Nitzan Rosenfeld; Florent Mouliere; Chandra S Chilamakuri; James A Morris; Aadhitthya Vijayaraghavan; Katrin Heider; Dineika Chandrananda; Wendy N Cooper; Davina Gale; Javier Garcia-Corbacho; Simon Pacey; Richard D Baird Journal: Genome Res Date: 2021-12-20 Impact factor: 9.043
Authors: Joaquim S L Vong; Peiyong Jiang; Suk-Hang Cheng; Wing-Shan Lee; Jason C H Tsang; Tak-Yeung Leung; K C Allen Chan; Rossa W K Chiu; Y M Dennis Lo Journal: Prenat Diagn Date: 2019-01-10 Impact factor: 3.050