Pyrite cloning: a single tube and programmed reaction cloning with restriction enzymes.

ABSTRACT:

Background

Insertion of engineered DNA fragments into bacterial vectors is the foundation of recombinant DNA technology, yet existing methods are still laborious, require many steps, depend on specific vector configuration, or require expensive reagents.

Results

We have developed a method, called "Pyrite" cloning that combines the traditional restriction enzyme digestion and ligation reaction in a single tube and uses a programmed thermocycler reaction, allowing rapid and flexible cloning in a single tube. After the Pyrite reaction and transformation, approximately 50% colonies contain the expected insert, which can be easily and quickly determined by colony PCR or blue-white colony screening. We also demonstrated that Pyrite cloning can be applied for different cloning purp

SUBMITTER: Fischer MD 

PROVIDER: S-EPMC6192209 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature