Production of closed linear DNA using a palindromic sequence

摘要

A primer for the amplification of a DNA template comprising a protelomerase target sequence, particularly for production of closed linear DNA, which primer is capable of specifically binding to a palindromic sequence within a protelomerase target sequence and priming amplification in both directions.

主权项

1. A process which is:
(I) a process for production of linear double stranded deoxyribonucleic acid (DNA) covalently closed at both ends by hairpin loops comprising: (a) amplifying by rolling circle amplification a DNA template comprising at least one protelomerase target sequence in the presence of at least one species of primer which binds specifically to a palindromic sequence within the at least one protelomerase target sequence and primes amplification in both directions, to produce a double stranded product, each strand comprising multiple protelomerase target sequences, wherein the amplification occurs in the absence of random primers; and (b) producing linear double stranded DNA covalently closed at both ends by hairpin loops by contacting amplified DNA comprising multiple protelomerase target sequences produced in (a) with at least one protelomerase under suitable conditions; (II) a process for making a pharmaceutical composition comprising a linear double stranded DNA covalently closed at both ends by hairpin loops, said process comprising carrying out a process according to (I), and formulating the resulting linear double stranded DNA covalently closed at both ends by hairpin loops with a pharmaceutically acceptable carrier or excipient, wherein (I) and (II) occur in an in vitro cell-free environment.