动物造模 z-bio 2022-11-08 481

　　Objective: To explore the feasibility of using Cas9 injection in mice after in vitro fertilization freeze-thaw.

　　Methods: Fresh C57BL/6J mouse eggs were frozen by cryopreservation tube method (EFS20/40) after IVF, and prokaryotic embryos (single-cell embryos) and 2-cell embryos were frozen the next day. The recovery and survival rates of frozen-thawed prokaryotic embryos and 2-cell embryos were compared. Partial frozen-thawed prokaryotic embryos and fresh prokaryotic embryos were simultaneously injected with Cas9 and diluted solution cytoplasm and cultured to 2 cells, and the survival rate and 2-cell development rate after injection were compared respectively.

　　Results: After freezing and thawing, the recovery rate of B6 mouse prokaryotic embryos was 92.5% and the survival rate was 92.8%. The recovery rate of 2-cell embryos was 90.5% and the survival rate was 95.8%. There was no significant difference between the two groups (P>0.05). ). The survival rate of fresh prokaryotic embryos after Cas9 injection was 92.7%, the blank group injection group was 97.5%, the survival rate of frozen-thawed prokaryotic embryos after Cas9 injection was 82.6%, and the blank group was 92%. There was significant (P<0.05), and="" other="" groups="" had="" no="" significant="" difference="" p="">0.05). There was no significant difference in the development rate of 2-cell embryos after injection (P>0.05).

　　Conclusion: Freeze-thawed prokaryotic embryos can be used for Cas9 microinjection.