Introducing precise genetic modifications into human 3PN embryos by CRISPR/Cas-mediated genome editing

Area of Science:

• Genetics
• Molecular Biology
• Bioengineering

Background:

• CRISPR/Cas is a powerful genome engineering tool.
• Its application in modifying various species is well-established.
• Precise genetic modification in human embryos requires careful evaluation.

Purpose of the Study:

• To evaluate CRISPR/Cas technology for precise genetic modifications in early human embryos.
• To establish principles for genome engineering in human embryos.

Main Methods:

• Human 3PN zygotes were injected with Cas9 mRNA and guide RNA (gRNA).
• Donor DNA (oligo or dsDonor) was co-injected with Cas9 mRNA and gRNA into embryos.
• Specific concentrations of mRNA, gRNA, and donor DNA were used.

Main Results:

• Successfully introduced the CCR5Δ32 allele into early human 3PN embryos using CRISPR/Cas.
• Observed incomplete control over other alleles at the targeted locus, resulting in wild-type or indel mutations.
• Demonstrated the potential and limitations of CRISPR/Cas for precise allele introduction.

Conclusions:

• Technical challenges in precise genome editing of human embryos remain.
• Implications for developing therapeutic treatments for genetic disorders exist.
• Advocates for rigorous global evaluation and discussion before human germline genome editing applications.