真核基因组DNA BamHI Hindlll BamHI Hindlll (5) 5) Sall Digest with Hindlll Digest with Sall 5P、 5'P、 cos ori ampr ori ampi2cos☐ Hindlll San BamHis'P San BamHI Hindlll 5'P San Hindlll BAP or CIP BAP or CIP cos ori amp ori amp☐cos☐ Sall BamHI BamHI Hindlll Digest with BamHl Digest with BamHI Isolate large fragment Isolate small fragment 5P、 cos■ ori amp Sall BamHl 5'P BamHI Hindlll Ligate to 35-45 kb fragments of eukaryotic DNA obtained by partial digestion with Mbol (or Sau3Al) 25