Abstract: Provided is a method capable of replicating or amplifying circular DNA, and particularly, long-chain circular DNA, in a cell-free system. Specifically, provided is a method for suppressing generation of a DNA multimer as a by-product, when circular DNA having a replication origin sequence (origin of chromosome (oriC)) is replicated or amplified by using the following enzyme groups: (1) a first enzyme group that catalyzes replication of circular DNA; (2) a second enzyme group that catalyzes an Okazaki fragment maturation and synthesizes two sister circular DNAs constituting a catenane; and (3) a third enzyme group that catalyzes a separation of two sister circular DNAs. Moreover, also provided is a method comprising introducing oriC into circular DNA by using a transposon.

Abstract: Provided is a method capable of simply and exponentially amplifying circular DNA, and particularly, long-chain circular DNA, in a cell-free system. Specifically, provided herein is a method for amplifying circular DNA which comprises mixing circular DNA having a replication origin sequence (origin of chromosome (oriC)) with a reaction solution comprising: a first enzyme group that catalyzes replication of circular DNA; a second enzyme group that catalyzes an Okazaki fragment maturation and synthesizes two sister circular DNAs constituting a catenane; a third enzyme group that catalyzes a separation of two sister circular DNAs; and also, a buffer, NTP, dNTP, a magnesium ion source, and an alkali metal ion source, to form a reaction mixture, which is then reacted.

Abstract: Provided is a method capable of replicating or amplifying circular DNA, and particularly, long-chain circular DNA, in a cell-free system. Specifically, provided is a method for suppressing generation of a DNA multimer as a by-product, when circular DNA having a replication origin sequence (origin of chromosome (oriC)) is replicated or amplified by using the following enzyme groups: (1) a first enzyme group that catalyzes replication of circular DNA; (2) a second enzyme group that catalyzes an Okazaki fragment maturation and synthesizes two sister circular DNAs constituting a catenane; and (3) a third enzyme group that catalyzes a separation of two sister circular DNAs. Moreover, also provided is a method comprising introducing oriC into circular DNA by using a transposon.