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. 2004 Feb 3;32(2):e23. doi: 10.1093/nar/gnh018

Figure 1.

Figure 1

Construction of DNA fragments containing an unmatched bulge. Two complementary synthetic oligonucleotides were designed to contain a specific mismatch or bulge after annealing to generate the middle part (MP). Two products, one containing an AvaI site and the other an Eco24I site, were PCR amplified from pBR322. The PCR products were digested with the corresponding enzymes, generating fragments with asymmetrical overhangs. Due to the overhang asymmetry, only a 298 bp tripartite DNA fragment (LEP–MP–REP) could be formed in a tripartite ligation (38). MP = middle part; LEP = left end part; REP = right end part. Gray tags represent overhangs.