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Nucleic Acids Res. Jul 1, 1996; 24(13): 2483–2487.
PMCID: PMC145980

A new class of genome rare cutters.


Although significant efforts have been directed at developing efficient techniques for rare and super rare genome cutting, only limited success has been achieved. Here we propose a new approach to solve this problem. We demonstrate that peptide nucleic acid 'clamps' (bis-PNAs) bind strongly and sequence specifically to short homopyrimidine sites on lambda and yeast genomic DNAs. Such binding efficiently shields methylation/restriction sites which overlap with the bis-PNA binding sites from enzymatic methylation. After removing the bis-PNA, the genomic DNAs are quantitatively cleaved by restriction enzymes into a limited number of pieces of lengths from several hundred kbp to several Mbp. By combining various bis-PNAs with different methylation/restriction enzyme pairs, a huge new class of genome rare cutters can be created. These cutters cover the range of recognition specificities where very few, if any, cutters are now available.

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Selected References

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  • Schwartz DC, Cantor CR. Separation of yeast chromosome-sized DNAs by pulsed field gradient gel electrophoresis. Cell. 1984 May;37(1):67–75. [PubMed]
  • Carle GF, Frank M, Olson MV. Electrophoretic separations of large DNA molecules by periodic inversion of the electric field. Science. 1986 Apr 4;232(4746):65–68. [PubMed]
  • Schwartz DC, Li X, Hernandez LI, Ramnarain SP, Huff EJ, Wang YK. Ordered restriction maps of Saccharomyces cerevisiae chromosomes constructed by optical mapping. Science. 1993 Oct 1;262(5130):110–114. [PubMed]
  • Wang YK, Huff EJ, Schwartz DC. Optical mapping of site-directed cleavages on single DNA molecules by the RecA-assisted restriction endonuclease technique. Proc Natl Acad Sci U S A. 1995 Jan 3;92(1):165–169. [PMC free article] [PubMed]
  • Murray AW, Szostak JW. Construction of artificial chromosomes in yeast. Nature. 1983 Sep 15;305(5931):189–193. [PubMed]
  • Burke DT, Carle GF, Olson MV. Cloning of large segments of exogenous DNA into yeast by means of artificial chromosome vectors. Science. 1987 May 15;236(4803):806–812. [PubMed]
  • Bhagwat AS. Restriction enzymes: properties and use. Methods Enzymol. 1992;216:199–224. [PubMed]
  • Perler FB, Davis EO, Dean GE, Gimble FS, Jack WE, Neff N, Noren CJ, Thorner J, Belfort M. Protein splicing elements: inteins and exteins--a definition of terms and recommended nomenclature. Nucleic Acids Res. 1994 Apr 11;22(7):1125–1127. [PMC free article] [PubMed]
  • Dujon B, Belfort M, Butow RA, Jacq C, Lemieux C, Perlman PS, Vogt VM. Mobile introns: definition of terms and recommended nomenclature. Gene. 1989 Oct 15;82(1):115–118. [PubMed]
  • Koob M. Conferring new cleavage specificities of restriction endonucleases. Methods Enzymol. 1992;216:321–329. [PubMed]
  • Koob M, Szybalski W. Cleaving yeast and Escherichia coli genomes at a single site. Science. 1990 Oct 12;250(4978):271–273. [PubMed]
  • Strobel SA, Dervan PB. Single-site enzymatic cleavage of yeast genomic DNA mediated by triple helix formation. Nature. 1991 Mar 14;350(6314):172–174. [PubMed]
  • Ferrin LJ, Camerini-Otero RD. Selective cleavage of human DNA: RecA-assisted restriction endonuclease (RARE) cleavage. Science. 1991 Dec 6;254(5037):1494–1497. [PubMed]
  • Koob M, Burkiewicz A, Kur J, Szybalski W. RecA-AC: single-site cleavage of plasmids and chromosomes at any predetermined restriction site. Nucleic Acids Res. 1992 Nov 11;20(21):5831–5836. [PMC free article] [PubMed]
  • Kur J, Koob M, Burkiewicz A, Szybalski W. A novel method for converting common restriction enzymes into rare cutters: integration host factor-mediated Achilles' cleavage (IHF-AC). Gene. 1992 Jan 2;110(1):1–7. [PubMed]
  • Gnirke A, Iadonato SP, Kwok PY, Olson MV. Physical calibration of yeast artificial chromosome contig maps by RecA-assisted restriction endonuclease (RARE) cleavage. Genomics. 1994 Nov 15;24(2):199–210. [PubMed]
  • Ferrin LJ, Camerini-Otero RD. Long-range mapping of gaps and telomeres with RecA-assisted restriction endonuclease (RARE) cleavage. Nat Genet. 1994 Apr;6(4):379–383. [PubMed]
  • Veselkov AG, Demidov VV, Frank-Kamenetskii MD, Nielsen PE. PNA as a rare genome-cutter. Nature. 1996 Jan 18;379(6562):214–214. [PubMed]
  • Nielsen PE, Egholm M, Berg RH, Buchardt O. Sequence-selective recognition of DNA by strand displacement with a thymine-substituted polyamide. Science. 1991 Dec 6;254(5037):1497–1500. [PubMed]
  • Cherny DY, Belotserkovskii BP, Frank-Kamenetskii MD, Egholm M, Buchardt O, Berg RH, Nielsen PE. DNA unwinding upon strand-displacement binding of a thymine-substituted polyamide to double-stranded DNA. Proc Natl Acad Sci U S A. 1993 Mar 1;90(5):1667–1670. [PMC free article] [PubMed]
  • Demidov VV, Yavnilovich MV, Belotserkovskii BP, Frank-Kamenetskii MD, Nielsen PE. Kinetics and mechanism of polyamide ("peptide") nucleic acid binding to duplex DNA. Proc Natl Acad Sci U S A. 1995 Mar 28;92(7):2637–2641. [PMC free article] [PubMed]
  • Nielsen PE, Egholm M, Berg RH, Buchardt O. Sequence specific inhibition of DNA restriction enzyme cleavage by PNA. Nucleic Acids Res. 1993 Jan 25;21(2):197–200. [PMC free article] [PubMed]
  • Betts L, Josey JA, Veal JM, Jordan SR. A nucleic acid triple helix formed by a peptide nucleic acid-DNA complex. Science. 1995 Dec 15;270(5243):1838–1841. [PubMed]
  • Egholm M, Christensen L, Dueholm KL, Buchardt O, Coull J, Nielsen PE. Efficient pH-independent sequence-specific DNA binding by pseudoisocytosine-containing bis-PNA. Nucleic Acids Res. 1995 Jan 25;23(2):217–222. [PMC free article] [PubMed]
  • Christensen L, Fitzpatrick R, Gildea B, Petersen KH, Hansen HF, Koch T, Egholm M, Buchardt O, Nielsen PE, Coull J, et al. Solid-phase synthesis of peptide nucleic acids. J Pept Sci. 1995 May-Jun;1(3):175–183. [PubMed]
  • Piña B, Berger S, Marcus GA, Silverman N, Agapite J, Guarente L. ADA3: a gene, identified by resistance to GAL4-VP16, with properties similar to and different from those of ADA2. Mol Cell Biol. 1993 Oct;13(10):5981–5989. [PMC free article] [PubMed]
  • Egholm M, Buchardt O, Christensen L, Behrens C, Freier SM, Driver DA, Berg RH, Kim SK, Norden B, Nielsen PE. PNA hybridizes to complementary oligonucleotides obeying the Watson-Crick hydrogen-bonding rules. Nature. 1993 Oct 7;365(6446):566–568. [PubMed]

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