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Botany JoVE (Open Access) Citable · DOI

Embryo Microinjection and Transplantation Technique for Nasonia vitripennis Genome Manipulation

DOI: 10.3791/56990-v
What you'll learn
  • ✓ Perform pre-blastoderm embryo collection and alignment in Nasonia vitripennis
  • ✓ Execute microinjection of embryos for genome modification
  • ✓ Transplant injected G0 embryos onto pre-stung host females
  • ✓ Generate heritable CRISPR/Cas9 modifications in N. vitripennis
Protocol

Microinjection of Nasonia vitripennis embryos is an essential method for generating heritable genome modifications. Described here is a detailed procedure for microinjection and transplantation of Nasonia vitripennis embryos, which will greatly facilitate future genome manipulation in this organism.

Difficulty
advanced
Total time
~3–5 days (colony rearing to viable offspring screening)
Model organism
Nasonia vitripennis
Biosafety
BSL-1

Steps

1
Establish and maintain N. vitripennis colony

Rear Nasonia vitripennis colony under controlled conditions to ensure consistent supply of mated females and healthy host specimens for embryo work.

▶ 00:47
2
Collect and align pre-blastoderm stage embryos

Extract early-stage embryos from ovipositor deposits, remove chorion, and position them in alignment for microinjection into the anterior end.

▶ 01:40
3
Microinject genome-modifying reagents into embryos

Deliver sgRNA and Cas9 solution into aligned embryos using precision microinjection to introduce heritable genomic modifications.

▶ 04:13
4
Transplant injected embryos onto pre-stung host

Transfer viable injected G0 embryos into the ovarian cavity of a pre-stung host female wasp to allow development and germline transmission.

▶ 05:58
5
Verify genome modifications in G0 and G1 progeny

Screen offspring for sgRNA and Cas9 integration and assess heritability of induced genomic changes across generations.

▶ 07:15
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