Highlights • Deficiency of zygotic genome activation gene Zscan5b causes chromosomal anomalies • Zscan5b binds to linker histone H1 and protects chromosomal structure • Irradiated Zscan5b-deficient ESCs show significantly increased DNA stress markers • Zscan5b-deficient ESCs develop small choriocarcinomas and embryonal carcinomas Summary Zygotic genome activation (ZGA) begins after fertilization and is essential for establishing pluripotency and genome stability.