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Evaluation of extraction methods for methylated cell-free fetal DNA from maternal plasma

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Abstract

Purpose

Recently, fetal placenta-specific epigenetic regions (FSERs) have been identified for quantification of cell-free fetal DNA (cff-DNA) for non-invasive prenatal testing (NIPT). The aim of this study was to evaluate the efficiencies of a column-based kit and magnetic bead-based kit for quantification of methylated FSERs from maternal plasma.

Methods

Maternal plasma was extracted from normal pregnant women within the gestational age of 10~13 weeks (n = 24). Total cell-free DNA (cf-DNA) was extracted using a column-based kit and magnetic bead-based kit from the plasma of the same pregnant woman, respectively. Methylated FSERs were enriched from the extracted total cf-DNA using a methyl-CpG-binding domain-based protein method. The four FSERs were simultaneously quantified by multiplex real-time polymerase chain reaction.

Results

Methylated FSERs were detected in all samples extracted from both kits. However, the amplification of FSERs showed significant differences in the extraction efficiency of methylated FSERs between the two extraction methods. The Ct values of methylated FSERs extracted using the column-based kit were significantly lower than those obtained using the magnetic bead-based kit (P < 0.001 for all FSERs). The quantity of methylated FSERs was significantly higher for extracted DNA using the column-based kit than that extracted using the magnetic bead-based kit (P < 0.001 for all FSERs). Time and cost for the process of extraction were similar for the column kit and magnetic bead-based kit.

Conclusions

Our findings demonstrate that the column-based kit was more effective than the magnetic bead-based kit for isolation of methylated FSERs from maternal plasma as assessed by FSER detection.

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Acknowledgments

We thank our staff who took the time and effort to participate in this study: Da Eun Lee, Ju Yeon Park, Do Jin Kim, Shin Young Kim, Hyun Jin Kim, Yeon Woo Lee, Ah Rum Oh, Shin Young Lee, So Min Seo, Hwa Jin Choi, and Kyoung Mee Han.

Funding

This research was supported by a grant (HI16C0628) from the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health and Welfare, Republic of Korea.

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Authors and Affiliations

  1. Laboratory of Medical Genetics, Medical Research Institute, Cheil General Hospital and Women’s Healthcare Center, Seoul, South Korea

    Ji Hyae Lim, Bom Yi Lee, Jin Woo Kim, So Yeon Park & Hyun Mee Ryu

  2. Department of Obstetrics and Gynecology, Cheil General Hospital and Women’s Healthcare Center, Dankook University College of Medicine, 1-19, Mookjung-dong, Chung-gu, Seoul, 100-380, South Korea

    You Jung Han, Jin Hoon Chung, Min Hyoung Kim, Dong Wook Kwak & Hyun Mee Ryu

  3. Genes Laboratory, Molecular Diagnostic Institute, Gyeonggi-do, South Korea

    Hee Back Choi

Authors
  1. Ji Hyae Lim
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  2. Bom Yi Lee
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  3. Jin Woo Kim
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  4. You Jung Han
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  5. Jin Hoon Chung
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  6. Min Hyoung Kim
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  7. Dong Wook Kwak
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  8. So Yeon Park
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  9. Hee Back Choi
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  10. Hyun Mee Ryu
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Corresponding author

Correspondence to Hyun Mee Ryu.

Ethics declarations

Appropriate ethical approval for this study was obtained from the ethics committee of the institutional review board at Cheil General Hospital, Korea (CGH-IRB-2016-5). All participants have agreed the written informed consents before collection of blood samples.

Conflict of interest

The authors declare that they have no competing interests.

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Lim, J.H., Lee, B.Y., Kim, J.W. et al. Evaluation of extraction methods for methylated cell-free fetal DNA from maternal plasma. J Assist Reprod Genet 35, 637–641 (2018). https://doi.org/10.1007/s10815-018-1114-8

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  • DOI: https://doi.org/10.1007/s10815-018-1114-8

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