Scheme for multiply-primed rolling circle amplification. Oligonucleotide primers complementary to the amplification target circle are hybridized to the circle. The 3′ ends of the DNA strands are indicated by arrowheads to show the polarity of polymerization. Thickened lines indicate the location of the original primer sequences within the product strands. The addition of DNA polymerase and deoxynucleoside triphosphates (dNTPs) to the primed circle results in the extension of each primer, and displacement of each newly synthesized strand results from elongation of the primer behind it. Secondary priming events can subsequently occur on the displaced product strands of the initial rolling circle amplification step. It is possible that the last intermediate shown in the figure makes only a small contribution to the overall yield of amplified DNA.
