Abstract
Approaches to direct solid phase sequencing of genomic and plasmid DNA have been developed using magnetic beads, coated with streptavidin, as solid support. The DNA is immobilized through selective incorporation of biotin into one of the strands. A single stranded template, suitable for sequencing, is obtained through strand-specific elution. Using this concept, in vitro amplified plasmid DNA and chromosomal DNA were sequenced directly from single colonies. The solid phase approach ensures that the amplification and the sequencing reactions can be performed under optimal conditions. The system was found to be suitable for sequencing using both isotope- and fluorescent-labelled primers.
MeSH Terms
Base Sequence
DNA, Bacterial/genetics,isolation & purification
DNA-Directed DNA Polymerase
Enterotoxins/genetics
Escherichia coli/genetics
Gene Amplification
Genes, Bacterial
Genetic Techniques
Molecular Sequence Data
Plasmids
Staphylococcus aureus/genetics
Templates, Genetic
Chemicals
DNA, Bacterial
Enterotoxins
enterotoxin A, Staphylococcal
DNA-Directed DNA Polymerase
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Hultman T
Department of Biochemistry and Biotechnology, Royal Institute of Technology, Stockholm, Sweden.
Ståhl S
Hornes E
Uhlén M
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