Codon-optimized FLP (FLPo) was PCR amplified from pDIRE and inserted into pCS2+ using restriction enzyme digest cloning. To remove the LG tag, Flp mRNA was synthesized from linearized pCS2-FLPo using the mMESSAGE mMACHINE SP6 kit, and the resulting mRNA was injected into single-cell embryos.