Quick identification and localization of CpG islands in large genomic fragments by partial digestion with Hpa II and Hha I.
More than 50% of mammalian genes are associated with CpG islands and thus they serve as a good gene marker. We have devised a simple method to scan large pieces of native or cloned genomic DNA for CpG islands. The method is based on the presence of multiple Hpa II and Hha I sites in CpG islands, at a frequency 30 times higher than in the rest of the genome. The steps include complete digestion of DNA with a rare-cutting restriction endonuclease (to produce large fragments with defined ends), partial digestion with Hpa II and Hha I, and subsequent Southern hybridization with an end probe. This identifies a CpG island as a cluster of sub-bands and, based on their electrophoretic mobility, one can immediately locate the island relative to the ends. For many vectors, universal probes flanking the cloning site are available, enabling the simultaneous analysis of a large number of samples. We demonstrated the usefulness of the method by analyzing known CpG islands in native genomic DNA and lambda, cosmid and P1 clones, and by isolating two novel transcribed islands from anonymous cosmid clones. Our method is quick, inexpensive, and can detect CpG islands with few or even no rare-cutter sites.
Kato
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R
Division of Disease Genes, Institute of Genetic Information, Kyushu University, Fukuoka, Japan.
Sasaki
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H
D000818:Animals; D010582:Bacteriophage lambda / Q000235:genetics; D003001:Cloning, Molecular; D003360:Cosmids / Q000235:genetics; D018899:CpG Islands / Q000235:genetics*; D019175:DNA Methylation; D019003:Deoxyribonuclease HpaII / Q000378:metabolism*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism*; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D015183:Restriction Mapping; D014158:Transcription, Genetic
D019003:Deoxyribonuclease HpaII; D015252:Deoxyribonucleases, Type II Site-Specific; C032229:GANTC-specific type II deoxyribonucleases