DNA In Maternal Plasma Research Articles

Noninvasive Prenatal Test by Cell-Free Fetal DNA in Maternal Plasma: Current Progress and Prospective Clinical Applications

Context: Prenatal testing aims to identify fetal chromosomal and genetic disorders prior to delivery. Current invasive procedures such as amniocentesis and chorionic villus sampling (CVS) pose a risk to mother and fetus and such diagnostic procedures are available only to high-risk pregnancies, which limits aneuploidy detection rate. The identification of cell-free fetal DNA (cffDNA) in maternal circulation has made noninvasive prenatal testing (NIPT) possible. This review seeks to highlight the necessity of investing in NIPT and briefly summarizes the technical aspects of the NIPT and application of this method in clinical practice. Evidence Acquisition: PubMed, OVID, SCOPUS, and the Cochrane database were searched for relevant articles published between 1995 and 2014, using appropriate keywords including prenatal screening, noninvasive testing, prenatal diagnosis, cell free fetal DNA, maternal circulation, chromosomal aneuploidies, trisomy, and sex determination. Results were restricted to systematic reviews, randomized clinical trials, meta-analysis, and observational studies. Results: The importance of prenatal diagnosis and risks associated with current invasive techniques makes NIPT research morally and commercially beneficial. The outstanding advantages of NIPT over current prenatal diagnosis techniques include increasing detection rate, enabling earlier diagnosis, and eliminating iatrogenic fetal loss and risk to the mother due to invasive procedures. At present, two major techniques for isolating cffDNA, namely digital PCR and massively parallel sequencing (MPS), have enabled the successful implementation of NIPT into clinical practice such as fetal sex determination, RhD genotyping, and fetal chromosomal aneuploidy detection. Conclusions: The advent of new NIPT using cffDNA has been regarded as a revolution in prenatal testing and has attracted significant commercial interest in the field. It is not overoptimistic to predict that NIPT will supplement or replace existing screening and diagnostic tools.

Maternal plasma DNA testing: experience of women counseled at a prenatal diagnosis center.

To evaluate the early introduction of circulating cell-free (ccf) DNA testing in a prenatal diagnosis center serving a statewide population. A retrospective chart review of patients at high aneuploidy risk counseled during the two 10-week periods that documents indication, risk, maternal age, insurance coverage, decisions, and reasoning behind that decision. Among the 299 included women, indication was advanced maternal age (17% with and 56% without an additional indication), positive serum screen (15%), and abnormal ultrasound (12%). Uptake increased from 10% to 17%, as did patient awareness of the test (4% to 14%). Women with lower copayments were more likely to complete testing (23% vs. 5%, p<0.001). Most women completing testing (75%) wanted to avoid an invasive procedure, while those declining cited testing would not change anything (47%), preferred diagnostic testing (16%), negative follow-up testing (20%), and cost/insurance issues (9%). One of 42 tests was positive (trisomy 21). Individual patient follow-up allows us to document ccfDNA-related patient decision-making. Nearly half of the women did not want further testing and one in seven preferred immediate diagnostic testing. Patient costs were a barrier to testing that, if avoided, could increase test uptake by 50% or more.

Israeli Society of Medical Genetics NIPT Committee Opinion 072013: Non-Invasive Prenatal Testing of Cell-Free DNA in Maternal Plasma for Detection of Fetal Aneuploidy

Non-invasive prenatal testing (NIPT) of cell-free fetal DNA in maternal plasma is a novel approach, designed for detecting common aneuploidies in the fetus. The Israeli Society of Medical Geneticists (ISMG) supports its use according to the guidelines stated herein. The clinical data collected thus far indicate that NIPT is highly sensitive in detecting trisomies 21 and 18, and fairly sensitive in detecting trisomy 13 and sex chromosome aneuploidies. Because false-positive results may occur, an abnormal result must be validated by invasive prenatal testing. At this juncture, NIPT does not replace existing prenatal screening tests for Down syndrome, as these are relatively inexpensive and cost-effective. Nonetheless, NIPT may be offered to women considered to be at high risk for fetal chromosomal abnormalities as early as 10 weeks of gestation. The ISMG states that NIPT should be an informed patient choice, and that pretest counseling regarding the limitations of NIPT is warranted. Women at high risk for genetic disorders not detected by NIPT should be referred for genetic counseling. A normal test result may be conveyed by a relevant healthcare provider, while an abnormal result should be discussed during a formal genetic consultation session.

Noninvasive prenatal testing by maternal plasma DNA analysis: Current practice and future applications

Prenatal screening of fetal chromosomal aneuploidies and some common genetic diseases is an integral part of antenatal care. Definitive prenatal diagnosis is conventionally achieved by the sampling of fetal genetic material by amniocentesis or chorionic villus sampling. Due to the invasiveness of those procedures, they are associated with a 1 in 200 chance of fetal miscarriage. Hence, researchers have been exploring noninvasive ways to sample fetal genetic material. The presence of cell-free DNA released by the fetus into the circulation of its mother was demonstrated in 1997. Circulating fetal DNA is therefore obtainable through the collection of a blood sample from the pregnant woman without posing any physical harm to the fetus. By analyzing this source of fetal genetic material, researchers have succeeded in developing DNA-based noninvasive tests for the assessment of Down syndrome and single gene diseases. Since the end of 2011, tests for the noninvasive assessment of chromosomal aneuploidies have become commercially available in parts of the world. Recommendations from professional groups have since been made regarding how these tests could be incorporated into the framework of existing prenatal screening programs. More recently, cell-free circulating fetal DNA analysis have been shown to be applicable to the deciphering of the fetal molecular karyotype, genome and methylome. It is envisioned that an increasing number of the noninvasive prenatal tests will become clinically available. The ethical, social and legal implications of the introduction of some of these tests would need to be discussed in the context of different cultures, societal values and the legal framework.

Maternal smoking does not affect the amount of cell-free fetal DNA in maternal plasma during the 1st trimester of pregnancy

CffDNA, from 344 non-smoking, 38 smoking and 33 ex-smoking pregnant women at 11 + 0–13 + 6 gestational weeks, was extracted and quantified by the multicopy DYS14, as the fetal DNA marker and using the quantitative real-time PCR 7300 detection system. The smoking habit was based on maternal self-report, confirmed by cotinine levels and male fetuses were verified by phenotype at birth. The genders of newborns were compared with DYS14-cffDNA analysis, achieving a 100% diagnostic accuracy of the test. A total of 177 non-smokers, 18 smokers and 22 ex-smoker pregnancies with male fetuses were identified by the cffDNA concentration. Results showed that smoking status was not associated with different amounts of DYS14-cffDNA (p = 0.159), suggesting the possibility of offering cffDNA testing to all pregnant women, even if they are active smokers or ex-smokers, and the test can be unadjusted for smoking status.

Detection of fetal sex chromosome aneuploidy by massively parallel sequencing of maternal plasma DNA: initial experience in a Chinese hospital

To evaluate the performance of a massively parallel sequencing (MPS)-based test in detecting fetal sex chromosome aneuploidy (SCA) and to present a comprehensive clinical counseling protocol for SCA-positive patients. This was a retrospective study in a large patient cohort of 5950 singleton pregnancies which underwent MPS-based testing as a prenatal screening test for trisomies 21, 18 and 13, with X and Y chromosomes as secondary findings, in Southwest Hospital in China. MPS-based SCA-positive women were offered the choice of knowing whether their SCA results were positive and those who did commenced a two-stage post-test clinical counseling protocol. In Stage 1, general information about SCA was given, and women were given the option of invasive testing for confirmation of findings; in Stage 2, those who had chosen to undergo invasive testing were informed about the specific SCA affecting their fetus and their management options. Thirty-three cases were classified as SCA-positive by MPS-based testing. After Stage 1 of the two-stage post-test clinical counseling session, 33 (100%) of these pregnant women chose to know the screening test results, and 25 (75.76%) underwent an invasive diagnostic procedure and karyotype analysis, in one of whom karyotyping failed. In thirteen cases, karyotyping confirmed the MPS-based test results (two X0 cases, seven XXX cases, three XXY cases and one XYY case), giving a positive predictive value of 54.17% (13/24 cases confirmed by karyotyping). After post-test clinical counseling session Stage 2, seven women chose to terminate the pregnancy: one X0 case, two XXX cases, the three XXY cases and the single XYY case. Six women decided to continue with pregnancy: one X0 case and five XXX cases. Our study showed the feasibility of clinical application of the MPS-based test in the non-invasive detection of fetal SCA. Together with a two-stage post-test clinical counseling protocol, it leads to a well-informed decision-making procedure.

Noninvasive prenatal testing for fetal trisomy 9 mosaicism by maternal plasma DNA sequencing

Abstract Maternal plasma DNA sequencing based noninvasive prenatal testing (NIPT) has been proven to be highly accurate in the detection of trisomy 21, 18, 13, X and Y, however, few reports have been made on its detection efficiency of rare complex aneuploidies. Here, we report a case of fetal trisomy 9 mosaicism identified by using NIPT, which may provide useful information for the further integration of NIPT into prenatal screening and diagnosis practice.

Fetal Aneuploidy Detection by Cell-Free DNA Sequencing for Multiple Pregnancies and Quality Issues with Vanishing Twins.

Non-invasive prenatal testing (NIPT) by random massively parallel sequencing of maternal plasma DNA for multiple pregnancies is a promising new option for prenatal care since conventional non-invasive screening for fetal trisomies 21, 18 and 13 has limitations and invasive diagnostic methods bear a higher risk for procedure related fetal losses in the case of multiple gestations compared to singletons. In this study, in a retrospective blinded analysis of stored twin samples, all 16 samples have been determined correctly, with four trisomy 21 positive and 12 trisomy negative samples. In the prospective part of the study, 40 blood samples from women with multiple pregnancies have been analyzed (two triplets and 38 twins), with two correctly identified trisomy 21 cases, confirmed by karyotyping. The remaining 38 samples, including the two triplet pregnancies, had trisomy negative results. However, NIPT is also prone to quality issues in case of multiple gestations: the minimum total amount of cell-free fetal DNA must be higher to reach a comparable sensitivity and vanishing twins may cause results that do not represent the genetics of the living sibling, as described in two case reports.

Prenatal diagnosis of fetal chromosome aneuploidy by massively parallel genomic sequencing

To perform non-invasive prenatal diagnosis (NIPD) of chromosome aneuploidy by detecting free DNA in maternal peripheral plasma by massively parallel genomic sequencing and determine the feasibility of detecting cell-free fetal DNA in chromosomal copy. The plasma samples from 1 264 gravidas were collected from February 2012 to July 2013 at our center. Those pregnant women with a gestational age of 13 to 33 weeks having high risks in serological screening or aged over 35 years or abnormal fetus on ultrasonography. The peripheral venous blood samples were drawn from pregnant women and plasma DNA was extracted for preparing a sequencing library. By using Illumina HiSeq2000, high-throughput sequencing was performed. Amnocentesis and karyotypic analysis were conducted for positive cases and those with negative results were followed up. By massively parallel genomic sequencing, 20 of them showed positive results. By the standard of chromosomal karyotypic analysis, there were 21 positive cases of 13 trisomy and the diagnostic accordance rate was 100%. Among 18 positive cases of 6 trisomy, there were amniocentesis (n = 5, except for one dead fetus), trisomy 18 (n = 4) and normal karyotype with placenta chimera (n = 1). One positive case of 13 trisomy was in accordance with karyotypic analysis. Massively parallel sequencing for NIPD may be used as further screening for pregnant women with high risks of serological screening. And this technology is highly consistent with karyotypic analysis in terms of sensitivity and specificity.

Genome-wide normalized score: a novel algorithm to detect fetal trisomy 21 during non-invasive prenatal testing

Non-invasive prenatal testing for fetal trisomy 21 (T21) by massively parallel shotgun sequencing (MPSS) is available for clinical use but its efficacy is limited by several factors, e.g. the proportion of cell-free fetal DNA in maternal plasma and sequencing depth. Existing algorithms discard DNA reads from the chromosomes for which testing is not being performed (i.e. those other than chromosome 21) and are thus more susceptible to diluted fetal DNA and limited sequencing depth. We aimed to describe and evaluate a novel algorithm for aneuploidy detection (genome-wide normalized score (GWNS)), which normalizes read counts by the proportions of DNA fragments from chromosome 21 in normal controls. We assessed the GWNS approach by comparison with two existing algorithms, i.e. Z-score and normalized chromosome value (NCV), using theoretical approximations and computer simulations in a set of 86 cases (64 euploid and 22 T21 cases). We then validated GWNS by studying an expanded set of clinical samples (n = 208). Finally, dilution experiments were undertaken to compare performance of the three algorithms (Z-score, NCV, GWNS) when fetal DNA concentration was low. At fixed levels of significance and power, GWNS required a smaller fetal DNA proportion and fewer total MPSS reads compared to Z-score or NCV. In dilution experiments, GWNS also outperformed the other two methods by reaching the correct diagnosis with the lowest range of fetal DNA concentrations (GWNS, 3.83-4.75%; Z-score, 4.75-5.22%; NCV, 6.47-8.58%). Our results demonstrate that GWNS is comparable to Z-score and NCV methods regarding the performance of detecting fetal T21. Dilution experiments suggest that GWNS may perform better than the other methods when fetal fraction is low.

Comparison of the performance of Ion Torrent chips in noninvasive prenatal trisomy detection.

Semiconductor high-throughput sequencing, represented by Ion Torrent PGM/Proton, proves to be feasible in the noninvasive prenatal diagnosis of fetal aneuploidies. It is commendable that, with less data and relevant cost also, an accurate result can be achieved owing to the high sensitivity and specificity of such kind of technology. We conducted a comparative analysis of the performance of four different Ion chips in detecting fetal chromosomal aneuploidies. Eight maternal plasma DNA samples, including four pregnancies with normal fetuses and four with trisomy 21 fetuses, were sequenced on Ion Torrent 314/316/318/PI chips, respectively. Results such as read mapped ratio, correlation coefficient and phred quality score were calculated and parallelly compared. All samples were correctly classified even with low-throughput chip, and, among the four chips, the 316 chip had the highest read mapped ratio, correlation coefficient, mean read length and phred quality score. All chips were well consistent with each other. Our results showed that all Ion chips are applicable in noninvasive prenatal fetal aneuploidy diagnosis. We recommend researchers or clinicians to use the appropriate chip with barcoding technology on the basis of the sample number.

Apert Syndrome in the Era of Prenatal Diagnosis

ABSTRACT Apert syndrome is a rare autosomal dominant disorder characterized by severe craniosynostosis, hypoplastic mediofacial structures and symmetric syndactyly. The syndactyly involves both cutaneous and osseous structures of the upper and lower limbs. Apart from skeletal anomalies, affected fetuses may often present central nervous system malformations and various visceral malformations (congenital heart defects, genitourinary anomalies, choanal stenosis, tracheal abnormalities, diaphragmatic hernia and omphalocele). Mental retardation is frequently encountered, as are a large variety of craniofacial and neurocognitive phenotypes. The incidence of Apert syndrome is approximately one in 65,000 newborns, representing 4.5% of all craniosynostosis cases. The first cases of Apert syndrome were diagnosed by ultrasound and fetoscopy in the third trimester of pregnancy. With the introduction of three-dimensional (3D) ultrasonography, identification of fetal changes characteristic of Apert syndrome is possible in the second trimester of pregnancy. Diagnosis confirmation requires fetal DNA analysis by fibroblast growth factor receptor 2 gene amplification (FGFR2) followed by sequencing and digestion with restriction enzymes. Recently, noninvasive prenatal diagnosis was reported using free fetal DNA analysis with cell-free fetal DNA (cffDNA) in maternal plasma by Polymerase Chain Reaction (PCR) assay and enzymatic digestion with restriction enzymes. As the syndrome has rarely been reported in Romania, we present the case of a fetus diagnosed with Apert syndrome in the second trimester of pregnancy. Early ultrasound detection of suggestive signs for this syndrome (craniosynostosis, syndactyly) was followed by confirmation of the diagnosis by prenatal fetal DNA analysis. How to cite this article Stamatian F, Kovacs T, Militaru M, Caracostea G. Apert Syndrome in the Era of Prenatal Diagnosis. Donald School J Ultrasound Obstet Gynecol 2014;8(2):222-225.

Non-Invasive Prenatal Testing Using Cell Free DNA in Maternal Plasma: Recent Developments and Future Prospects.

Recent advances in molecular genetic technologies have facilitated non-invasive prenatal testing (NIPT) through the analysis of cell-free fetal DNA in maternal plasma. NIPT can be used to identify monogenic disorders including the identification of autosomal recessive disorders where the maternally inherited mutation needs to be identified in the presence of an excess of maternal DNA that contains the same mutation. In the future, simultaneous screening for multiple monogenic disorders is anticipated. Several NIPT methods have been developed to screen for trisomy. These have been shown to be effective for fetal trisomy 21, 18 and 13. Although the testing has been extended to sex chromosome aneuploidy, robust estimates of the efficacy are not yet available and maternal mosaicism for gain or loss of an X-chromosome needs to be considered. Using methods based on the analysis of single nucleotide polymorphisms, diandric triploidy can be identified. NIPT is being developed to identify a number of microdeletion syndromes including α-globin gene deletion. NIPT is a profoundly important development in prenatal care that is substantially advancing the individual patient and public health benefits achieved through conventional prenatal screening and diagnosis.

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.

Noninvasive prenatal testing for autosomal recessive conditions by maternal plasma sequencing in a case of congenital deafness.

The goals of our study were to develop a noninvasive prenatal test for autosomal recessive monogenic conditions and to prove its overall feasibility and potential for clinical integration. We recruited a pregnant woman and her spouse, who had a proband child suffering from congenital deafness, and obtained the target-region sequencing data from a semicustom array that used genomic and maternal plasma DNA from three generations of this family. A haplotype-assisted strategy was developed to detect whether the fetus inherited the pathogenic mutations in the causative gene, GJB2. The parental haplotype was constructed using a trio strategy through two different processes, namely, the grandparent-assisted haplotype phasing process and the proband-assisted haplotype phasing process. The fetal haplotype was deduced afterward based on both the maternal plasma sequencing data and the parental haplotype. The accuracy levels of paternal and maternal haplotypes obtained by grandparent-assisted haplotype phasing were 99.01 and 97.36%, respectively, and the proband-assisted haplotype phasing process yielded slightly lower accuracies of 98.73 and 96.79%, respectively. Fetal inheritance of the pathogenic gene was deduced correctly in both processes. Our study indicates that the strategy of haplotype-based noninvasive prenatal testing for monogenic conditions has potential applications in clinical practice.

Noninvasive prenatal diagnosis of common aneuploidies by semiconductor sequencing.

Massively parallel sequencing (MPS) of cell-free fetal DNA from maternal plasma has revolutionized our ability to perform noninvasive prenatal diagnosis. This approach avoids the risk of fetal loss associated with more invasive diagnostic procedures. The present study developed an effective method for noninvasive prenatal diagnosis of common chromosomal aneuploidies using a benchtop semiconductor sequencing platform (SSP), which relies on the MPS platform but offers advantages over existing noninvasive screening techniques. A total of 2,275 pregnant subjects was included in the study; of these, 515 subjects who had full karyotyping results were used in a retrospective analysis, and 1,760 subjects without karyotyping were analyzed in a prospective study. In the retrospective study, all 55 fetal trisomy 21 cases were identified using the SSP with a sensitivity and specificity of 99.94% and 99.46%, respectively. The SSP also detected 16 trisomy 18 cases with 100% sensitivity and 99.24% specificity and 3 trisomy 13 cases with 100% sensitivity and 100% specificity. Furthermore, 15 fetuses with sex chromosome aneuploidies (10 45,X, 2 47,XYY, 2 47,XXX, and 1 47,XXY) were detected. In the prospective study, nine fetuses with trisomy 21, three with trisomy 18, three with trisomy 13, and one with 45,X were detected. To our knowledge, this is the first large-scale clinical study to systematically identify chromosomal aneuploidies based on cell-free fetal DNA using the SSP and provides an effective strategy for large-scale noninvasive screening for chromosomal aneuploidies in a clinical setting.

Noninvasive prenatal testing for microdeletion syndromes and expanded trisomies: proceed with caution.

The identification of circulating cell-free fetal DNA in maternal plasma has led to the introduction of noninvasive prenatal tests with high sensitivity and high specificity for common aneuploidies (trisomy 13, trisomy 18, trisomy 21). A new expanded noninvasive prenatal testing panel that includes five microdeletion syndromes (22q11 deletion syndrome, cri-du-chat [5p minus], Prader Willi or Angelman syndrome, 1p36 deletion syndrome) and two aneuploidies usually associated with nonviable pregnancies (trisomy 16 and trisomy 22) is now available. This expanded panel will be performed unless an opt-out box is checked. Because these disorders are so rare, the positive predictive value is expected to be low. As with all new screening tests and technologies, the expanded panel should be appropriately studied before it is widely adopted.

Epigenome-wide DNA methylation assay reveals placental epigenetic markers for noninvasive fetal single-nucleotide polymorphism genotyping in maternal plasma.

The use of DNA methylation difference between maternal blood cell and fetal (placental) DNA is one of the main areas of interest for the development of fetal epigenetics markers in maternal plasma. We employed a methylation array (HumanMethylation450 array, Illumina, Inc.) to identify novel biomarkers that are specially hypermethylated in placental DNA versus maternal blood cells in a genome-wide basis. Validation by bisulfite genomic sequencing was performed and the priority was given to potential targets that harbor differential methylated CpG sites overlapped with at least two methylation-sensitive restriction enzyme (MSRE) recognizing sites, as well as one polymorphic single-nucleotide polymorphism (SNP), within a short DNA stretch. Three candidate regions of PSMB8, SKI, and CHST11 gene were selected for developing a preliminary polymerase chain reaction assay with MSRE digestion of maternal plasma DNA. SNP genotypes were confirmed by direct sequencing. We identified 2944 and 5218 fetal-specific hypermethylated CpG sites in the first- and third-trimester placenta, respectively, of which 2613 were overlapped, suggesting a consistency of differential methylation during the whole pregnancy. The array results were confirmed by bisulfite genomic sequencing. The preliminary tests in maternal plasma showed that postdigestion hypermathylated versions of these candidate molecules were detectable only in pregnant women. We further revealed that methylated targets in maternal plasma possessed the fetal SNP genotypes. The present studies systematically identified hypermethylated sites in fetal tissues and preliminarily demonstrated that some of the fetal epigenetic markers that contain informative SNPs have great potential for noninvasive fetal genetic diagnosis.

Accurate Fetal Chromosome Dosage Determination by Shotgun Sequencing of Maternal Plasma DNA without PCR Amplification during Library Preparation

To the Editor: Nextgen sequencing technologies have enabled noninvasive prenatal testing of fetal aneuploidies by shotgun sequencing of maternal plasma DNA during pregnancy (1). A statistically significant change (e.g., using z -score statistics) in the number of reads mapped to a chromosome of interest (such as chromosome 21) is used to identify fetal aneuploidy. The accuracy of the test is largely dependent on 2 factors: the variability (SD) of the percentage of reads mapped to the chromosome of interest among euploidy samples and the absolute difference between the sample being tested and the mean of the euploidy samples. We speculated that the PCR enrichment step in the library preparation before cluster generation and sequencing contributes to the variation in chromosome dosage determination. Additionally, with 3 ng of DNA (typically obtainable from about 2 mL of maternal plasma) at an average size of 150 bp per DNA fragment, around 20 billion DNA fragments were available for sequencing, suggesting PCR amplification may not be necessary. We thus modified the current protocol for library preparation from maternal plasma DNA by removing the PCR enrichment step. Maternal plasma samples of about 2.5–3.2 mL were obtained with informed consent. DNA preparation and sequencing library preparation were performed as previously described (2), up to the step for adaptor ligation and purification. The purified ligation products were then equally divided into 2 …

Scientific Impact Paper No. 15: Non‐invasive Prenatal Testing for Chromosomal Abnormality using Maternal Plasma DNA

Scientific Impact Paper No. 15: Non‐invasive Prenatal Testing for Chromosomal Abnormality using Maternal Plasma DNA