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Pronuclear removal of tripronuclear zygotes can establish heteroparental normal karyotypic human embryonic stem cells

  • Stem Cell Biology
  • Published:
Journal of Assisted Reproduction and Genetics Aims and scope Submit manuscript

Abstract

Purpose

This study aimed to derive heteroparental normal karyotypic human embryonic stem cells (hESCs) from microsurgically corrected tripronuclear (3PN) zygotes.

Methods

After sequential culture for 5–6 days, embryos developed from microsurgically corrected 3PN zygotes were analyzed by fluorescence in situ hybridization (FISH) using probes for chromosomes 17, X and Y. Intact 3PN zygotes from clinical in vitro fertilization (IVF) cycles were cultured as the control group. The inner cell mass (ICM) of blastocysts that developed from microsurgically corrected 3PN zygotes was used to derive hESC lines, and the stem cell characteristics of these lines were evaluated. G-banding analysis was adopted to identify the karyotype of the hESC line, and the heteroparental inheritance of the hESC line was analyzed by DNA fingerprinting analysis.

Results

The blastocyst formation rate (13.5 %) of the microsurgically corrected 3PN zygotes was significantly higher (P < 0.05) than that of intact 3PN zygotes (8.7 %). The diploid rate of the blastocysts (55.0 %) was significantly higher (P < 0.05) than that of the arrested cleavage-stage embryos (18.4 %) in microsurgically corrected 3PN zygotes. The triploid rate of the microsurgically corrected 3PN zygotes (5.7 %) was significantly lower (P < 0.01) than that of intact 3PN zygotes (19.4 %). Furthermore, we established one heteroparental normal karyotypic hESC line from the microsurgically corrected tripronuclear zygotes.

Conclusions

Pronuclear removal can effectively remove the surplus chromosome set of 3PN zygotes. A combination of pronuclear removal and blastocyst culture enables the selection of diploidized blastocysts from which heteroparental normal karyotypic hESC lines can be derived.

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Acknowledgments

This work was supported by grants from the Major State Basic Research Development Program of China (no. 2012CB944901) and the National Science Foundation of China (no. 81222007), and by the Program for New Century Excellent Talents in University.

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Author notes

    Authors and Affiliations

    1. The Second Affiliated Hospital of South China University, 30th Jie-Fang Road, Hengyang, Hunan, 421001, China

      Hong-Qing Liao

    2. Institute of Reproductive and Stem Cell Engineering, Central South University, Changsha, Hunan, 410078, China

      Qi OuYang, Shuo-Ping Zhang, Guang-Xiu Lu & Ge Lin

    3. National Engineering and Research Center of Human Stem Cell, Changsha, Hunan, 410078, China

      Qi OuYang, Guang-Xiu Lu & Ge Lin

    4. CITIC-Xiangya Reproductive and Genetic Hospital, Changsha, Hunan, 410078, China

      Shuo-Ping Zhang, De-Hua Cheng, Guang-Xiu Lu & Ge Lin

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    1. Hong-Qing Liao
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    Correspondence to Hong-Qing Liao or Ge Lin.

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    Capsule

    Pronuclear removal can effectively remove the surplus chromosome set of 3PN zygotes. A combination of pronuclear removal and blastocyst culture enables the selection of diploidized blastocysts from which heteroparental normal karyotypic hESC lines can be derived.

    Hong-Qing Liao and Qi OuYang contributed equally to this work.

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    Liao, HQ., OuYang, Q., Zhang, SP. et al. Pronuclear removal of tripronuclear zygotes can establish heteroparental normal karyotypic human embryonic stem cells. J Assist Reprod Genet 33, 255–263 (2016). https://doi.org/10.1007/s10815-015-0634-8

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    • DOI: https://doi.org/10.1007/s10815-015-0634-8

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