Size-based molecular diagnostics using plasma DNA for noninvasive prenatal testing.
Noninvasive prenatal testing using fetal DNA in maternal plasma is an actively researched area. The current generation of tests using massively parallel sequencing is based on counting plasma DNA sequences originating from different genomic regions. In this study, we explored a different approach that is based on the use of DNA fragment size as a diagnostic parameter. This approach is dependent on the fact that circulating fetal DNA molecules are generally shorter than the corresponding maternal DNA molecules. First, we performed plasma DNA size analysis using paired-end massively parallel sequencing and microchip-based capillary electrophoresis. We demonstrated that the fetal DNA fraction in maternal plasma could be deduced from the overall size distribution of maternal plasma DNA. The fetal DNA fraction is a critical parameter affecting the accuracy of noninvasive prenatal testing using maternal plasma DNA. Second, we showed that fetal chromosomal aneuploidy could be detected by observing an aberrant proportion of short fragments from an aneuploid chromosome in the paired-end sequencing data. Using this approach, we detected fetal trisomy 21 and trisomy 18 with 100% sensitivity (T21: 36/36; T18: 27/27) and 100% specificity (non-T21: 88/88; non-T18: 97/97). For trisomy 13, the sensitivity and specificity were 95.2% (20/21) and 99% (102/103), respectively. For monosomy X, the sensitivity and specificity were both 100% (10/10 and 8/8). Thus, this study establishes the principle of size-based molecular diagnostics using plasma DNA. This approach has potential applications beyond noninvasive prenatal testing to areas such as oncology and transplantation monitoring.
Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China;Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, NT, Hong Kong SAR, China;
Stephanie C Y
SC
Yu
Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China;Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, NT, Hong Kong SAR, China;
K C Allen
KC
Chan
Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China;Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, NT, Hong Kong SAR, China;
Yama W L
YW
Zheng
Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China;Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, NT, Hong Kong SAR, China;
Peiyong
P
Jiang
Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China;Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, NT, Hong Kong SAR, China;
Gary J W
GJ
Liao
Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China;Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, NT, Hong Kong SAR, China;
Hao
H
Sun
Harris Birthright Research Centre for Fetal Medicine, King's College Hospital, London SE5 9RS, United Kingdom;
Ranjit
R
Akolekar
Department of Obstetrics and Gynaecology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, NT, Hong Kong SAR, China; and.
Tak Y
TY
Leung
Department of Clinical Chemistry, VU University Medical Center, 1081 HV, Amsterdam, The Netherlands.
Attie T J I
AT
Go
Department of Clinical Chemistry, VU University Medical Center, 1081 HV, Amsterdam, The Netherlands.
John M G
JM
van Vugt
Harris Birthright Research Centre for Fetal Medicine, King's College Hospital, London SE5 9RS, United Kingdom;
Ryoko
R
Minekawa
Department of Clinical Chemistry, VU University Medical Center, 1081 HV, Amsterdam, The Netherlands.
Cees B M
CB
Oudejans
Harris Birthright Research Centre for Fetal Medicine, King's College Hospital, London SE5 9RS, United Kingdom;
Kypros H
KH
Nicolaides
Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China;Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, NT, Hong Kong SAR, China;
Rossa W K
RW
Chiu
Centre for Research into Circulating Fetal Nucleic Acids, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China;Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, NT, Hong Kong SAR, China; loym@cuhk.edu.hk.
Y M Dennis
YM
Lo
D025063:Chromosome Disorders / Q000175:diagnosis / Q000235:genetics; D002882:Chromosomes, Human, Pair 13 / Q000235:genetics; D002887:Chromosomes, Human, Pair 18 / Q000235:genetics; D041321:Chromosomes, Human, X / Q000235:genetics; D004247:DNA / Q000097:blood / Q000737:chemistry / Q000235:genetics*; D004314:Down Syndrome / Q000175:diagnosis / Q000235:genetics; D019075:Electrophoresis, Capillary / Q000379:methods; D005260:Female; D005315:Fetal Diseases / Q000097:blood / Q000175:diagnosis / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D009006:Monosomy / Q000175:diagnosis / Q000235:genetics; D057089:Pathology, Molecular / Q000379:methods*; D011247:Pregnancy; D011296:Prenatal Diagnosis / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014314:Trisomy / Q000175:diagnosis / Q000235:genetics; D000073839:Trisomy 13 Syndrome; D000073842:Trisomy 18 Syndrome
D004247:DNA
Down syndrome; Turner syndrome; fetal aneuploidy; next-generation sequencing; size profiling