DNA Sequencing Directly from a Mixture Using Terminal-Base-Selective Primers

DNA cloning is often used to select and amplify one DNA species from a mixture. However, the cloning process is complex and labor-intensive. We have developed a new two-step method for DNA sequencing directly from a mixture. The first is the introduction of a known oligonucleotide (common part) into the terminus of unknown DNA by ligation. The second is selective DNA sequencing using primers with two additional nucleotides at the 3 ' terminus in addition to the common part (terminal-base-selective primers). The primers work only for templates on which the primers perfectly hybridized. This method was found to be effective for the Hindlll digestion products of A phage. Key words: DNA sequencing; cloning; terminal-base-selective primer; DNA sequencing of mixture 1