In vitro production of large single-stranded templates for DNA sequencing.
A method has been developed for the preparation of large single-stranded DNA sequencing templates from primary cloning plasmids or cosmids. The method utilizes the separate action of T7 Gene 6 exonuclease and exonuclease III to generate large quantities of single-stranded template for each strand of a large-cloned fragment. Since the procedure is intended for use on primary clones, it avoids the time-consuming subcloning steps associated with most sequencing programs. The procedure also has the advantage of avoiding clone instability problems associated with subcloning in M13.
Straus
N A
NA
Department of Botany, University of Toronto, Ontario, Canada.
Zagursky
R J
RJ
D001483:Base Sequence; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D004274:DNA, Recombinant; D004277:DNA, Single-Stranded / Q000096:biosynthesis / Q000235:genetics*; D005090:Exodeoxyribonucleases / Q000378:metabolism; D008969:Molecular Sequence Data; D010957:Plasmids; D013698:Templates, Genetic
D004274:DNA, Recombinant; D004277:DNA, Single-Stranded; D005090:Exodeoxyribonucleases; C046264:gene b exonuclease; C020133:exodeoxyribonuclease III; D004262:DNA Restriction Enzymes