A new enzymatic route for production of long 5'-phosphorylated oligonucleotides using suicide cassettes and rolling circle DNA synthesis-6

Copyright information:Taken from "A new enzymatic route for production of long 5'-phosphorylated oligonucleotides using suicide cassettes and rolling circle DNA synthesis"http://www.biomedcentral.com/1472-6750/7/49BMC Biotechnology 2007;7():49-49.Published online 16 Aug 2007PMCID:PMC2040145.ization in the suicide cassette region. The DNA circle is closed by self-templated ligation. By the addition of a primer, complementary to part of the DNA sequence, dNTPs and a polymerase a rolling circle DNA synthesis can be initiated resulting in tandem repeats complementary to the templating circle. The suicide cassette can be removed from the oligonucleotide by cleavage with the Mly I restriction enzyme. The tandem repeats can be linearized without the loss of nucleotides using the Nt. Alw I nicking enzyme, resulting in monomers complementary to the initial DNA sequence. Self-templated hybridization (see step A). Self-templated ligation (see step B). Rolling circle DNA synthesis (see step C). Cleavage with Mly I (see step D). Nicking, using Nt. Alw I, resulting in monomers of the same polarity as the initial DNA sequence. (+) and (-) indicate the polarity of the oligonucleotide.