Inverse fusion PCR cloning.

Spiliotis, Markus (2012). Inverse fusion PCR cloning. PLoS ONE, 7(4), e35407. Public Library of Science 10.1371/journal.pone.0035407

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Inverse fusion PCR cloning (IFPC) is an easy, PCR based three-step cloning method that allows the seamless and directional insertion of PCR products into virtually all plasmids, this with a free choice of the insertion site. The PCR-derived inserts contain a vector-complementary 5'-end that allows a fusion with the vector by an overlap extension PCR, and the resulting amplified insert-vector fusions are then circularized by ligation prior transformation. A minimal amount of starting material is needed and experimental steps are reduced. Untreated circular plasmid, or alternatively bacteria containing the plasmid, can be used as templates for the insertion, and clean-up of the insert fragment is not urgently required. The whole cloning procedure can be performed within a minimal hands-on time and results in the generation of hundreds to ten-thousands of positive colonies, with a minimal background.

Item Type:

Journal Article (Original Article)

Division/Institute:

05 Veterinary Medicine > Department of Infectious Diseases and Pathobiology (DIP) > Institute of Parasitology

UniBE Contributor:

Spiliotis, Markus

Subjects:

600 Technology > 630 Agriculture

ISSN:

1932-6203

Publisher:

Public Library of Science

Language:

English

Submitter:

Andrea Stettler

Date Deposited:

16 Nov 2015 17:31

Last Modified:

16 Nov 2015 17:31

Publisher DOI:

10.1371/journal.pone.0035407

PubMed ID:

22530019

BORIS DOI:

10.7892/boris.73126

URI:

https://boris.unibe.ch/id/eprint/73126

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