Ficoll-400 discontinuous density gradient centrifugation was used to purify superficial yolk spheres under blastoderms of fertilized chicken eggs and the corresponding site of unfertilized eggs. Under microscope, the purified yolk spheres showed good morphology and no contamination from embronic cells was observed. The purified yolk spheres were lysed and digested with high concentration of proteinase K then extracted for long time with phenol. When subjected to agarose gel electrophoresis, the DNA showed a clear band. The recovery of DNA is approximately 10 ng per fertilized or unfertilized egg.