Workshop report on the extraction of foetal DNA from maternal plasma

Legler, Tobias J.; Liu, Zhong; Mavrou, Ariadni; Finning, Kirstin; Hromadníková, Ilona; Galbiati, Silvia; Meaney, Cathy; Hultén, Maj; Crea, Francesco; Olsson, Martin L.; Maddocks, Deborah G.; Huang, Dorothy Jane; Fisher, Sylvia Armstrong; Sprenger-Haussels, Markus; Soussan, Aïcha Ait; Schoot, C. Ellen van der

doi:10.1002/pd.1783

Cited by 112 publications

(81 citation statements)

References 22 publications

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“…First successful applications of the EZ1 system, which requires considerable less handson time of technical laboratory assistants than tradition column-based extraction systems [23], were published in the fi elds of microbiology [24] and forensic medicine [25,26] as early as in 2005. The EZ1 system was shown to be useful for the isolation of target DNA in low quantities, e.g., of fetal DNA in maternal plasma [21], or of DNA in partially degraded biological specimens [22]. Successful EZ1-based DNA extraction from formalin-fi xed, paraffi nembedded tissue samples [9], or otherwise long-term preserved tissue specimens [8] has been demonstrated.…”

Section: Discussionmentioning

confidence: 99%

Comparison of an automated nucleic acid extraction system with the column-based procedure

Frickmann¹,

Hinz²,

Hagen³

2015

European Journal of Microbiology and Immunology

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Here, we assessed the extraction efficiency of a deployable bench-top nucleic acid extractor EZ1 in comparison to the column-based approach with complex sample matrices.A total of 48 EDTA blood samples and 81 stool samples were extracted by EZ1 automated extraction and the column-based QIAamp DNA Mini Kit. Blood sample extractions were assessed by two real-time malaria PCRs, while stool samples were analyzed by six multiplex real-time PCR assays targeting bacterial, viral, and parasitic stool pathogens. Inhibition control PCR testing was performed as well.In total, 147 concordant and 13 discordant pathogen-specific PCR results were obtained. The latter comprised 11 positive results after column-based extraction only and two positive results after EZ1 extraction only. EZ1 extraction showed a higher frequency of inhibition. This phenomenon was, however, inconsistent for the different PCR schemes. In case of concordant PCR results, relevant differences of cycle threshold numbers for the compared extraction schemes were not observed.Switches from well-established column-based extraction to extraction with the automated EZ1 system do not lead to a relevantly reduced yield of target DNA when complex sample matrices are used. If sample inhibition is observed, column-based extraction from another sample aliquot may be considered.

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Section: Discussionmentioning

confidence: 99%

Comparison of an automated nucleic acid extraction system with the column-based procedure

Frickmann¹,

Hinz²,

Hagen³

2015

European Journal of Microbiology and Immunology

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“…28,29 Hence, for manual reference methods, it has been demonstrated that the QIAamp DSP Virus Kit might be the optimal tool because it extracts mainly small fragments of DNA, finally providing larger amounts of cffDNA. 30,31 Second, regarding false-positive results, it has been observed that a multicopy sequence, such as DYS14, is more sensitive, accurate, and efficient than the single-copy SRY in the assessment of cffDNA, which is particularly important early in the first trimester of pregnancy when the copy numbers of fetal DNA are low. 32 The drawback of the DYS14 sequence is that it has considerable homology to sequences other than the Y chromosome that could falsely classify female fetuses as male.…”

Section: Genetics In Medicine | Volume 14 | Number 1 | January 2012mentioning

confidence: 99%

Noninvasive fetal sex determination in maternal plasma: a prospective feasibility study

Fernández-Martínez¹,

Galindo

García-Burguillo

et al. 2012

Genetics in Medicine

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Purpose: To prospectively validate a protocol for noninvasive fetal sex determination in maternal plasma and demonstrate its applicability to clinical practice. methods: Peripheral blood from 404 pregnant women undergoing prenatal invasive testing was collected from 6 to 23 weeks of gestation. Real-time PCR was performed for the SRY gene and multicopy DYS14 marker sequence located within the TSPY gene by the TaqMan minor groove binder probe assay as a first-line test. Owing to a false-positive result, amplification of repetitive motifs of the DAZ gene region was also tested as a second-line test performed in the last 232 patients enrolled in our series. A diagnostic algorithm was designed using a combination of these three markers. Fetal gender determined by noninvasive prenatal diagnosis (NIPD) was compared with that diagnosed by quantitative fluorescent PCR after invasive testing or ultrasound.Results: A single false-positive result was obtained in the first 172 pregnancies. Reporting criteria were modified in the subsequent 232 pregnancies, giving an overall sensitivity and specificity of 100% (95% CI 99.8-100%) and 99.5% (95% CI 98

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“…In particular, RT-MSP of EF3 marker on bisulfite converted plasma DNA samples was carried out assuming that the mean yield of DNA extracted per 0.5 ml of plasma is 10 ng, of which 5% is of fetal origin, as suggested by Legler et al 44 and assuming that the yield of DNA after bisulfite conversion is 65%. To verify the fetal origin of the RT-MSP product from maternal plasma samples we amplified plasma DNA, the corresponding amounts of maternal blood cell DNA and CVS DNA as negative and positive controls respectively.…”

Section: Ef3 Discriminates Cffdna Versus Cfmdna In Maternal Plasma Samentioning

confidence: 99%

Differential DNA Methylation as a Tool for Noninvasive Prenatal Diagnosis (NIPD) of X Chromosome Aneuploidies

Ragione

Mastrovito

Campanile

et al. 2010

The Journal of Molecular Diagnostics

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The demographic tendency in industrial countries to delay childbearing , coupled with the maternal age effect in common chromosomal aneuploidies and the risk to the fetus of invasive prenatal diagnosis, are potent drivers for the development of strategies for noninvasive prenatal diagnosis. One breakthrough has been the discovery of differentially methylated cell-free fetal DNA in the maternal circulation. We describe novel bisulfite conversion-and methylationsensitive enzyme digestion DNA methylation-related approaches that we used to diagnose Turner syndrome from first trimester samples. We used an Xlinked marker , EF3 , and an autosomal marker, RASSF1A , to discriminate between placental and maternal blood cell DNA using real-time methylationspecific PCR after bisulfite conversion and real-time PCR after methylation-sensitive restriction digestion. By normalizing EF3 amplifications versus RASSF1A outputs , we were able to calculate sex chromosome/ autosome ratios in chorionic villus samples , thus permitting us to correctly diagnose Turner syndrome. The identification of this new marker coupled with the strategy outlined here may be instrumental in the development of an efficient , noninvasive method of diagnosis of sex chromosome aneuploidies in plasma

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Workshop report on the extraction of foetal DNA from maternal plasma

Abstract: This workshop initiated a standardization process for extraction of cff DNA in maternal plasma. The highest yield was obtained by the QIAamp DSP Virus Kit, a result that will be evaluated in more detail in future studies.

Cited by 112 publications

References 22 publications

Comparison of an automated nucleic acid extraction system with the column-based procedure

Comparison of an automated nucleic acid extraction system with the column-based procedure

Noninvasive fetal sex determination in maternal plasma: a prospective feasibility study

Differential DNA Methylation as a Tool for Noninvasive Prenatal Diagnosis (NIPD) of X Chromosome Aneuploidies

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