This study describes an optimized blastocyst complementation (IBC) strategy using C-CRISPR to facilitate the rapid screening of candidate genes and successfully generate rat forebrain tissue in mice via IBC. The *Hesx1* gene deficiency was identified as supporting the generation of functional rat forebrain tissue in mice. Xenogeneic rat forebrain tissues in adult mice were structurally and functionally intact. Comparative analyses revealed that rat forebrain tissues developed at the same pace as the mouse host but maintained rat-like transcriptome profiles. The chimeric rate of rat cells gradually decreased during mid-to-late prenatal development, indicating xenogeneic barriers. The study demonstrates that *Hesx1*^−/−^ mouse blastocysts provide a developmental niche suitable for the formation of functional rat forebrain tissues from donor rat embryonic stem cells (rESCs). The findings highlight the potential of the C-CRISPR-based IBC strategy to broaden the study and application of interspecies organogenesis, particularly in understanding brain development and cognitive function.This study describes an optimized blastocyst complementation (IBC) strategy using C-CRISPR to facilitate the rapid screening of candidate genes and successfully generate rat forebrain tissue in mice via IBC. The *Hesx1* gene deficiency was identified as supporting the generation of functional rat forebrain tissue in mice. Xenogeneic rat forebrain tissues in adult mice were structurally and functionally intact. Comparative analyses revealed that rat forebrain tissues developed at the same pace as the mouse host but maintained rat-like transcriptome profiles. The chimeric rate of rat cells gradually decreased during mid-to-late prenatal development, indicating xenogeneic barriers. The study demonstrates that *Hesx1*^−/−^ mouse blastocysts provide a developmental niche suitable for the formation of functional rat forebrain tissues from donor rat embryonic stem cells (rESCs). The findings highlight the potential of the C-CRISPR-based IBC strategy to broaden the study and application of interspecies organogenesis, particularly in understanding brain development and cognitive function.