Assembling multiple fragments: the Gibson Assembly

Luisana Avilan

doi:10.1007/978-1-0716-3004-4_4

Assembling multiple fragments: the Gibson Assembly

Luisana Avilan

Research output: Chapter in Book/Report/Conference proceeding › Chapter (peer-reviewed) › peer-review

Abstract

The Gibson Assembly is a popular method for molecular cloning which has been developed specifically to join several fragments together in a specific order, without the constraint of restriction enzyme sites. This method is based on the assembly of overlapping fragments, generally produced by PCR, and then combining them using three enzymes: a 5′ exonuclease, a DNA polymerase, and a DNA ligase, in an isothermal reaction. Here, we describe this method, including the design of primers for the generation of the overlapping fragments and the assembly; to this end, we provide an example involving joining two fragments in a single plasmid.

Original language	English
Title of host publication	DNA Manipulation and Analysis
Editors	Garry Scarlett
Place of Publication	New York
Publisher	Humana Press
Chapter	4
Pages	45-53
Number of pages	9
Edition	1st
ISBN (Electronic)	9781071630044
ISBN (Print)	9781071630037
DOIs	https://doi.org/10.1007/978-1-0716-3004-4_4
Publication status	Published - 1 Mar 2023

Publication series

Name	Methods in Molecular Biology
Volume	2633
ISSN (Print)	1064-3745
ISSN (Electronic)	1940-6029

Keywords

Gibson Assembly
cloning
overlapping primers

Access to Document

10.1007/978-1-0716-3004-4_4

Cite this

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ER -

Assembling multiple fragments: the Gibson Assembly

Abstract

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Keywords

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