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Use of Bisection to Reduce Mitochondrial DNA in the Bovine Oocyte
Published on: July 6, 2022
Interspecies somatic cell nuclear transfer is dependent on compatible mitochondrial DNA and reprogramming factors.
Yan Jiang1, Richard Kelly, Amy Peters
1Mitochondrial and Reproductive Genetics Group, Clinical Sciences Research Institute, Warwick Medical School, Coventry, United Kingdom.
Plos One
|May 11, 2011
Summary
Improving interspecies somatic cell nuclear transfer (iSCNT) requires compatible mitochondrial DNA (mtDNA). Supplementing with species-specific mtDNA and reprogramming factors significantly enhances blastocyst development in reconstructed embryos.
Area of Science:
- Reproductive biology
- Developmental biology
- Genetics
Background:
- Interspecies somatic cell nuclear transfer (iSCNT) faces challenges in reprogramming donor nuclei and mitochondrial DNA (mtDNA) compatibility.
- Early embryonic arrest and failure to express key pluripotency markers are common in iSCNT embryos.
- Mitochondrial DNA plays a crucial role in cellular energy production (ATP) and embryonic development.
Purpose of the Study:
- To investigate the impact of nuclear-cytoplasmic compatibility on embryonic development in a murine-porcine iSCNT model.
- To determine the effect of supplementing enucleated oocytes with species-compatible mtDNA and reprogramming factors on iSCNT outcomes.
- To analyze the mtDNA content and profile throughout preimplantation development in iSCNT embryos.
Main Methods:
- Murine fetal fibroblasts were transferred into enucleated porcine oocytes.
- Porcine oocytes were depleted of mtDNA using 2',3'-dideoxycytidine.
- Murine somatic cells and murine embryonic stem cell extract were transferred into mtDNA-depleted oocytes.
- Allele-specific PCR was used to quantify mtDNA content and profile.
- Blastocyst rates and Oct-4 expression were assessed.
Main Results:
- Standard iSCNT resulted in very low blastocyst rates (0.48%) and failed nuclear reprogramming.
- Murine mtDNA levels were extremely low (<0.001%) in standard iSCNT embryos by the blastocyst stage.
- Supplementation with murine mtDNA and ESC extract significantly increased blastocyst rates to 3.38% (P<0.01).
- Supplemented embryos showed significantly higher levels of murine mtDNA throughout development, reaching ~50% at later stages.
- Supplemented embryos exhibited mtDNA profiles similar to murine embryos.
Conclusions:
- Species-compatible mtDNA is critical for successful preimplantation development in iSCNT.
- Supplementing oocytes with donor species' mtDNA and reprogramming factors improves developmental competence.
- Addressing nuclear-cytoplasmic incompatibility, particularly mtDNA, is key to advancing iSCNT technologies.
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