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The presence of fetal DNA in maternal plasma has opened up fascinating possibilities for noninvasive prenatal testing [1,2]. Recently, there has been much interest inside the use of massively parallel sequencing (MPS) for analyzing circulating fetal DNA for prenatal testing purposes. Therefore, fetal trisomies 21, 13, 18 and selected sex chromosomal aneuploidies have been detected applying MPS on maternal plasma DNA [3?] and have been quickly introduced into clinical service. Apart from abnormalities on account of copy number adjustments involving a whole chromosome, it will be critical to evaluate whether the MPS-based evaluation of maternal plasma could be sensitive sufficient for detecting subchromosomal deletions or duplications. Within this regard, Peters et al reported the detection of a 4.2 Mb deletion on chromosome 12 within a maternal plasma sample obtained at the 35th week of gestation [8]. Jensen et al reported the detection of a three Mb deletion on chromosome 22 inPLOS A single | plosone.orgmaternal plasma samples obtained from two pregnant women in the 19th and 20th weeks of gestation [9]. Apart from the deleted region, Peters et al also performed statistical analysis on a further area on chromosome 12, as well as 20 nonoverlapping 4 Mb regions on chromosome 14 [8]. Jensen et al, on the other hand, only focused their statistical analysis on the deleted region on chromosome 22 [9]. Hence, from the data presented by Peters et al and Jensen et al, it truly is not clear when the approach would be robust sufficient for any genomewide survey of microdeletions or microduplications, or certainly for the noninvasive determination of a fetal karyotype. Lo et al reported that fetal single nucleotide polymorphisms (SNPs) is often genotyped within a genomewide scale employing maternal plasma DNA sequencing [10]. In specific, these investigators have demonstrated that SNP alleles and mutations for single gene disorders that happen to be inherited by a fetus from its mother is often elucidated by a process known as relative haplotype dosage evaluation [10]. Fan et al confirmed the robustness of relative haplotypeNoninvasive Prenatal Molecular Karyotypingdosage analysis and made use of this method to detect a ,two.Formula of 1420898-14-1 85 Mb deletion inherited by a fetus from its mother [11].Furan-2,4(3H,5H)-dione Price You’ll find two concerns for making use of this technique for the clinical implementation of noninvasive prenatal karyotyping.PMID:24631563 Very first, this approach demands maternal haplotyping to be performed which would call for further analytical steps [12,13] or pedigree analysis. Second, it can be unclear if this approach might be utilized to detect de novo subchromosomal deletion or duplication. We’ve not too long ago shown that through the use of shotgun MPS on the plasma DNA of cancer individuals, one particular could noninvasively get a `plasma karyotype’ of a cancer at 1 Mb resolution [14,15]. In this report, we sought to apply a comparable strategy for acquiring the prenatal molecular karyotypes of many fetuses by shotgun MPS of maternal plasma DNA.Sample Processing and DNA ExtractionPeripheral blood samples had been centrifuged at 1600 g for ten min at 4uC and the plasma portion was recentrifuged at 16000 g for 10 min at 4uC [17]. We extracted cell-free DNA from 1.8 to 8.four mL of maternal plasma using the QIAamp DSP DNA Blood Mini Kit (Qiagen) as described previously [3]. The extracted plasma DNA was quantified by a real-time PCR assay targeting the.
