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Cell Biology JoVE (Open Access) Citable · DOI

Methods for Precisely Localized Transfer of Cells or DNA into Early Postimplantation Mouse Embryos

DOI: 10.3791/53295-v
What you'll learn
  • ✓ Perform precise surgical dissection and ex vivo culture of postimplantation mouse embryos
  • ✓ Graft cultured cells into defined anatomical sites within early embryos
  • ✓ Deliver exogenous DNA via optimized electroporation using glass capillaries
  • ✓ Assess localized cell and DNA transfer outcomes in embryonic tissue
Protocol

We demonstrate a method for grafting cultured cells into defined sites of early mouse embryos to determine their in vivo potential. We also introduce an optimized electroporation method that uses glass capillaries of known diameter, allowing the precise delivery of exogenous DNA into a few cells in the embryos.

Difficulty
advanced
Total time
~4–6 hours per embryo (dissection, grafting/electroporation, imaging)
Model organism
Mouse (E7.5–E8.5 postimplantation embryos)
Biosafety
BSL-1

Steps

1
Dissect E7.5 or E8.5 postimplantation mouse embryos

Remove implanted embryos from uteri and prepare them for ex vivo culture. Carefully isolate embryos from maternal tissue and establish dissection conditions.

▶ 00:45
2
Graft cultured cells into embryonic tissue

Position cultured cells at defined anatomical sites within intact early embryos using micromanipulation. Monitor cell integration and survival during subsequent culture.

▶ 02:03
3
Prepare handmade electroporation capillaries and apparatus

Fabricate glass capillaries of known diameter and assemble the electroporation setup. Calibrate electrode spacing and electrical parameters for precise DNA delivery.

▶ 03:42
4
Deliver exogenous DNA via electroporation

Apply voltage pulses through positioned glass capillaries to introduce DNA into target embryonic cells. Verify pulse parameters and cell viability post-treatment.

▶ 05:19
5
Assess localized cell and DNA transfer outcomes

Examine grafted cells and transfected tissue regions by imaging. Document spatial distribution and expression patterns of transferred cells or reporter genes.

▶ 06:44
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