Dideoxy sequencing method using denatured plasmid templates.
作者： Masahira Hattori , Yoshiyuki Sakaki
DOI: 10.1016/0003-2697(86)90403-3
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摘要: Abstract The dideoxy sequencing method in which denatured plasmid DNA is used as a template was improved. simple and rapid: the recombinant extracted purified by rapid alkaline lysis followed ribonuclease treatment. then immediately with alkali subjected to reaction utilizing synthetic oligonucleotide primers. It takes only several hours from start of extraction end reaction. We examined each step procedure, points were found be crucial for making reproducible powerful: (i) should free RNA open circular (or linear) DNA; (ii) heptadecamer rather than pentadecamer recommended primer; (iii) done at 37°C or higher room temperature. enabled us determine sequence more thousand nucleotides single DNA.
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