Methods for nucleic acid manipulation
First Claim
1. A method of replication of a target nucleic acid sequence contained in a nucleic acid duplex comprising contacting said nucleic acid duplex with primers that are complementary to the flanking ends of said target sequence and with a polymerase, in the presence of bacteriophage T4 UvsX protein.
2 Assignments
0 Petitions
Accused Products
Abstract
A method for replicating and amplifying a target nucleic acid sequence is described. A method of the invention involves the formation of a recombination intermediate without the prior denaturing of a nucleic acid duplex through the use of a recombination factor. The recombination intermediate is treated with a high fidelity polymerase to permit the replication and amplification of the target nucleic acid sequence. In preferred embodiments, the polymerase comprises a polymerase holoenzyme. In further preferred embodiments, the recombination factor is bacteriophage T4 UvsX protein or homologs from other species, and the polymerase holoenzyme comprises a polymerase enzyme, a clamp protein and a clamp loader protein, derived from viral, bacteriophage, prokaryotic, archaebacterial, or eukaryotic systems.
26 Citations
12 Claims
- 1. A method of replication of a target nucleic acid sequence contained in a nucleic acid duplex comprising contacting said nucleic acid duplex with primers that are complementary to the flanking ends of said target sequence and with a polymerase, in the presence of bacteriophage T4 UvsX protein.
-
12. A method of replication of a target nucleic acid sequence contained in a nucleic acid duplex comprising contracting said nucleic acid duplex with primers that are complementary to the flanking ends of said target sequence and with a polymerase, in the presence of bacteriophage T4 UvsX protein at a temperature of 37°
- C.
Specification