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Nucleic Acids Res. Nov 11, 1989; 17(21): 8543–8551.
PMCID: PMC335026

A computer program for choosing optimal oligonucleotides for filter hybridization, sequencing and in vitro amplification of DNA.


A method is presented for choosing optimal oligodeoxyribonucleotides as probes for filter hybridization, primers for sequencing, or primers for DNA amplification. Three main factors that determine the quality of a probe are considered: stability of the duplex formed between the probe and target nucleic acid, specificity of the probe for the intended target sequence, and self-complementarity. DNA duplex stability calculations are based on the nearest-neighbor thermodynamic values determined by Breslauer et al. [Proc. Natl. Acad. Sci. U.S.A. (1986), 83: 3746]. Temperatures of duplex dissociation predicted by the method described here were within 0.4 degrees C of the values obtained experimentally for ten oligonucleotides. Calculations for specificity of the probe and its self-complementarity are based on a simple dynamic algorithm.

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Selected References

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  • Breslauer KJ, Frank R, Blöcker H, Marky LA. Predicting DNA duplex stability from the base sequence. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3746–3750. [PMC free article] [PubMed]
  • Freier SM, Kierzek R, Jaeger JA, Sugimoto N, Caruthers MH, Neilson T, Turner DH. Improved free-energy parameters for predictions of RNA duplex stability. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9373–9377. [PMC free article] [PubMed]
  • Rychlik W, Domier LL, Gardner PR, Hellmann GM, Rhoads RE. Amino acid sequence of the mRNA cap-binding protein from human tissues. Proc Natl Acad Sci U S A. 1987 Feb;84(4):945–949. [PMC free article] [PubMed]
  • Nussinov R, Jacobson AB. Fast algorithm for predicting the secondary structure of single-stranded RNA. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6309–6313. [PMC free article] [PubMed]
  • Zuker M, Stiegler P. Optimal computer folding of large RNA sequences using thermodynamics and auxiliary information. Nucleic Acids Res. 1981 Jan 10;9(1):133–148. [PMC free article] [PubMed]
  • Williams AL, Jr, Tinoco I., Jr A dynamic programming algorithm for finding alternative RNA secondary structures. Nucleic Acids Res. 1986 Jan 10;14(1):299–315. [PMC free article] [PubMed]

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