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DNA sequencing with direct blotting electrophoresis.
Author(s) -
Beck S.,
Pohl F.M.
Publication year - 1984
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1002/j.1460-2075.1984.tb02230.x
Subject(s) - dna , biology , gel electrophoresis , dna sequencing , nucleotide , southern blot , blot , microbiology and biotechnology , nucleic acid sequence , genetics , gene
A method for transferring the DNA molecules of sequencing reaction mixtures onto an immobilizing matrix during electrophoresis has been developed. A blotting membrane moves with constant speed across the end of a very short, denaturing gel and collects the molecules according to size. A constant distance between bands for molecules differing in length by one nucleotide is obtained over a large range (approximately 600 nucleotides with a 5% gel), simplifying the determination of DNA sequences considerably. Reliable sequences of 500 nucleotides can be read and sequence features up to greater than 1000 nucleotides are revealed in a single experiment. The sequencing of a potential Z‐DNA‐forming fragment from Escherichia coli DNA is given as an example and possible further developments are discussed.