The development of novel quantification assay for mitochondrial DNA heteroplasmy aimed at preimplantation genetic diagnosis of Leigh encephalopathy

Hiroto Tajima, Kou Sueoka, Sung Yung Moon, Akira Nakabayashi, Tomoyoshi Sakurai, Yukitaka Murakoshi, Hiroyoshi Watanabe, Soukichi Iwata, Tsuyoshi Hashiba, Shingo Kato, Yu Ichi Goto, Yasunori Yoshimura

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

Purpose: To perform preimplantation genetic diagnosis (PGD) of Leigh encephalopathy, we developed a rapid and reliable quantification assay for the percentage of T8993G mtDNA mutation and analyzed various specimens. Methods: We prepared the standard curve by measuring serial proportion of 8993T/G cloned plasmid DNA using real-time PCR, and measured (1) mutant DNA (known proportions by PCR-RFLP), (2) single lymphocytes from 46% mutant carrier, (3) 123 blastomeres from 20 abnormal embryos. Results: (1) These were within -5∼+6% error range, (2) mean 44.3%(11-70%), (3) Five embryos harbored T8993G mutation (4-22%). Embryos from same person indicated different degrees of heteroplasmy, and blastomeres from same embryo demonstrated limited dispersion of heteroplasmy (2-11%). Conclusions: (1) This method provides rapid and reliable PGD for Leigh encephalopathy. (2) The variable heteroplasmy with somatic mitosis was suggested. (3) T8993G mutation was existed in undeveloped embryo, and the bottleneck theory was supported. The limited heteroplasmy dispersion of blastomeres from same embryo also supported reliability of PGD for T8993G mutation.

Original languageEnglish
Pages (from-to)227-232
Number of pages6
JournalJournal of Assisted Reproduction and Genetics
Volume24
Issue number6
DOIs
Publication statusPublished - 2007 Jun

Fingerprint

Preimplantation Diagnosis
Brain Diseases
Mitochondrial DNA
Embryonic Structures
Blastomeres
Mutation
DNA
Mitosis
Restriction Fragment Length Polymorphisms
Real-Time Polymerase Chain Reaction
Plasmids
Lymphocytes
Polymerase Chain Reaction

Keywords

  • Heteroplasmy
  • Leigh encephalopathy
  • Mitochondrial DNA
  • Preimplantation genetic diagnosis
  • Real-time PCR

ASJC Scopus subject areas

  • Obstetrics and Gynaecology
  • Developmental Biology
  • Genetics
  • Reproductive Medicine

Cite this

The development of novel quantification assay for mitochondrial DNA heteroplasmy aimed at preimplantation genetic diagnosis of Leigh encephalopathy. / Tajima, Hiroto; Sueoka, Kou; Moon, Sung Yung; Nakabayashi, Akira; Sakurai, Tomoyoshi; Murakoshi, Yukitaka; Watanabe, Hiroyoshi; Iwata, Soukichi; Hashiba, Tsuyoshi; Kato, Shingo; Goto, Yu Ichi; Yoshimura, Yasunori.

In: Journal of Assisted Reproduction and Genetics, Vol. 24, No. 6, 06.2007, p. 227-232.

Research output: Contribution to journalArticle

Tajima, H, Sueoka, K, Moon, SY, Nakabayashi, A, Sakurai, T, Murakoshi, Y, Watanabe, H, Iwata, S, Hashiba, T, Kato, S, Goto, YI & Yoshimura, Y 2007, 'The development of novel quantification assay for mitochondrial DNA heteroplasmy aimed at preimplantation genetic diagnosis of Leigh encephalopathy', Journal of Assisted Reproduction and Genetics, vol. 24, no. 6, pp. 227-232. https://doi.org/10.1007/s10815-007-9114-0
Tajima, Hiroto ; Sueoka, Kou ; Moon, Sung Yung ; Nakabayashi, Akira ; Sakurai, Tomoyoshi ; Murakoshi, Yukitaka ; Watanabe, Hiroyoshi ; Iwata, Soukichi ; Hashiba, Tsuyoshi ; Kato, Shingo ; Goto, Yu Ichi ; Yoshimura, Yasunori. / The development of novel quantification assay for mitochondrial DNA heteroplasmy aimed at preimplantation genetic diagnosis of Leigh encephalopathy. In: Journal of Assisted Reproduction and Genetics. 2007 ; Vol. 24, No. 6. pp. 227-232.
@article{297a0997697d4c15a83c032a0cb8bf3e,
title = "The development of novel quantification assay for mitochondrial DNA heteroplasmy aimed at preimplantation genetic diagnosis of Leigh encephalopathy",
abstract = "Purpose: To perform preimplantation genetic diagnosis (PGD) of Leigh encephalopathy, we developed a rapid and reliable quantification assay for the percentage of T8993G mtDNA mutation and analyzed various specimens. Methods: We prepared the standard curve by measuring serial proportion of 8993T/G cloned plasmid DNA using real-time PCR, and measured (1) mutant DNA (known proportions by PCR-RFLP), (2) single lymphocytes from 46{\%} mutant carrier, (3) 123 blastomeres from 20 abnormal embryos. Results: (1) These were within -5∼+6{\%} error range, (2) mean 44.3{\%}(11-70{\%}), (3) Five embryos harbored T8993G mutation (4-22{\%}). Embryos from same person indicated different degrees of heteroplasmy, and blastomeres from same embryo demonstrated limited dispersion of heteroplasmy (2-11{\%}). Conclusions: (1) This method provides rapid and reliable PGD for Leigh encephalopathy. (2) The variable heteroplasmy with somatic mitosis was suggested. (3) T8993G mutation was existed in undeveloped embryo, and the bottleneck theory was supported. The limited heteroplasmy dispersion of blastomeres from same embryo also supported reliability of PGD for T8993G mutation.",
keywords = "Heteroplasmy, Leigh encephalopathy, Mitochondrial DNA, Preimplantation genetic diagnosis, Real-time PCR",
author = "Hiroto Tajima and Kou Sueoka and Moon, {Sung Yung} and Akira Nakabayashi and Tomoyoshi Sakurai and Yukitaka Murakoshi and Hiroyoshi Watanabe and Soukichi Iwata and Tsuyoshi Hashiba and Shingo Kato and Goto, {Yu Ichi} and Yasunori Yoshimura",
year = "2007",
month = "6",
doi = "10.1007/s10815-007-9114-0",
language = "English",
volume = "24",
pages = "227--232",
journal = "Journal of Assisted Reproduction and Genetics",
issn = "1058-0468",
publisher = "Springer New York",
number = "6",

}

TY - JOUR

T1 - The development of novel quantification assay for mitochondrial DNA heteroplasmy aimed at preimplantation genetic diagnosis of Leigh encephalopathy

AU - Tajima, Hiroto

AU - Sueoka, Kou

AU - Moon, Sung Yung

AU - Nakabayashi, Akira

AU - Sakurai, Tomoyoshi

AU - Murakoshi, Yukitaka

AU - Watanabe, Hiroyoshi

AU - Iwata, Soukichi

AU - Hashiba, Tsuyoshi

AU - Kato, Shingo

AU - Goto, Yu Ichi

AU - Yoshimura, Yasunori

PY - 2007/6

Y1 - 2007/6

N2 - Purpose: To perform preimplantation genetic diagnosis (PGD) of Leigh encephalopathy, we developed a rapid and reliable quantification assay for the percentage of T8993G mtDNA mutation and analyzed various specimens. Methods: We prepared the standard curve by measuring serial proportion of 8993T/G cloned plasmid DNA using real-time PCR, and measured (1) mutant DNA (known proportions by PCR-RFLP), (2) single lymphocytes from 46% mutant carrier, (3) 123 blastomeres from 20 abnormal embryos. Results: (1) These were within -5∼+6% error range, (2) mean 44.3%(11-70%), (3) Five embryos harbored T8993G mutation (4-22%). Embryos from same person indicated different degrees of heteroplasmy, and blastomeres from same embryo demonstrated limited dispersion of heteroplasmy (2-11%). Conclusions: (1) This method provides rapid and reliable PGD for Leigh encephalopathy. (2) The variable heteroplasmy with somatic mitosis was suggested. (3) T8993G mutation was existed in undeveloped embryo, and the bottleneck theory was supported. The limited heteroplasmy dispersion of blastomeres from same embryo also supported reliability of PGD for T8993G mutation.

AB - Purpose: To perform preimplantation genetic diagnosis (PGD) of Leigh encephalopathy, we developed a rapid and reliable quantification assay for the percentage of T8993G mtDNA mutation and analyzed various specimens. Methods: We prepared the standard curve by measuring serial proportion of 8993T/G cloned plasmid DNA using real-time PCR, and measured (1) mutant DNA (known proportions by PCR-RFLP), (2) single lymphocytes from 46% mutant carrier, (3) 123 blastomeres from 20 abnormal embryos. Results: (1) These were within -5∼+6% error range, (2) mean 44.3%(11-70%), (3) Five embryos harbored T8993G mutation (4-22%). Embryos from same person indicated different degrees of heteroplasmy, and blastomeres from same embryo demonstrated limited dispersion of heteroplasmy (2-11%). Conclusions: (1) This method provides rapid and reliable PGD for Leigh encephalopathy. (2) The variable heteroplasmy with somatic mitosis was suggested. (3) T8993G mutation was existed in undeveloped embryo, and the bottleneck theory was supported. The limited heteroplasmy dispersion of blastomeres from same embryo also supported reliability of PGD for T8993G mutation.

KW - Heteroplasmy

KW - Leigh encephalopathy

KW - Mitochondrial DNA

KW - Preimplantation genetic diagnosis

KW - Real-time PCR

UR - http://www.scopus.com/inward/record.url?scp=34547641390&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=34547641390&partnerID=8YFLogxK

U2 - 10.1007/s10815-007-9114-0

DO - 10.1007/s10815-007-9114-0

M3 - Article

C2 - 17342424

AN - SCOPUS:34547641390

VL - 24

SP - 227

EP - 232

JO - Journal of Assisted Reproduction and Genetics

JF - Journal of Assisted Reproduction and Genetics

SN - 1058-0468

IS - 6

ER -