Methodology

Protocol: Precision engineering of plant gene loci by homologous recombination cloning in Escherichia coli

Laura C Roden^1,3 , Berthold Göttgens² and Effie S Mutasa-Göttgens¹

¹Broom's Barn Research Station, Higham, Bury St Edmunds, Suffolk IP28 6NP, UK

²Cambridge Institute for Medical Research, Hills Road, Cambridge, CB2 2XY, UK

³Dept. Mol. & Cell Biol., UCT, Private Bag Rondebosch, 7701, Cape Town, South Africa

author email corresponding author email

Plant Methods 2005, 1:6doi:10.1186/1746-4811-1-6

Published:	29 September 2005

Abstract

Plant genome sequence data now provide opportunities to conduct molecular genetic studies at the level of the whole gene locus and above. Such studies will be greatly facilitated by adopting and developing further the new generation of genetic engineering tools, based on homologous recombination cloning in Escherichia coli, which are free from the constraints imposed by the availability of suitably positioned restriction sites. Here we describe the basis for homologous recombination cloning in E. coli, the available tools and resources, together with a protocol for long range cloning and manipulation of an Arabidopsis thaliana gene locus, to create constructs co-ordinately driven by locus-specific regulatory elements.