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Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate.

Proceedings of the National Academy of Sciences of the United States of America, 01 Aug 1979, 76(8):3683-3687
https://doi.org/10.1073/pnas.76.8.3683 PMID: 291033 PMCID: PMC383897

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Abstract 

We describe a technique for transferring electrophoretically separated bands of double-stranded DNA from agarose gels to diazobenzyloxymethyl-paper. Controlled cleavage of the DNA in situ by sequential treatment with dilute acid, which causes partial depurination, and dilute alkali, which causes cleavage and separation of the strands, allows the DNA to leave the gel rapidly and completely, with an efficiency independent of its size. Covalent attachment of DNA to paper prevents losses during subsequent hybridization and washing steps and allows a single paper to be reused many times. Ten percent dextran sulfate, originally found to accelerate DNA hybridization in solution by about 10-fold [J.G. Wetmur (1975) Biopolymers 14, 2517-2524], accelerates the rate of hybridization of randomly cleaved double-stranded DNA probes to immobilized nucleic acids by as much as 100-fold, without increasing the background significantly.

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Proc Natl Acad Sci U S A. 1979 Aug; 76(8): 3683–3687.
PMCID: PMC383897
PMID: 291033

Efficient transfer of large DNA fragments from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dextran sulfate.

G M Wahl, M Stern, and G R Stark
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Abstract

We describe a technique for transferring electrophoretically separated bands of double-stranded DNA from agarose gels to diazobenzyloxymethyl-paper. Controlled cleavage of the DNA in situ by sequential treatment with dilute acid, which causes partial depurination, and dilute alkali, which causes cleavage and separation of the strands, allows the DNA to leave the gel rapidly and completely, with an efficiency independent of its size. Covalent attachment of DNA to paper prevents losses during subsequent hybridization and washing steps and allows a single paper to be reused many times. Ten percent dextran sulfate, originally found to accelerate DNA hybridization in solution by about 10-fold [J.G. Wetmur (1975) Biopolymers 14, 2517-2524], accelerates the rate of hybridization of randomly cleaved double-stranded DNA probes to immobilized nucleic acids by as much as 100-fold, without increasing the background significantly.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
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