USER cloning is a fast and versatile method for engineering of plasmid DNA. We have developed an easy to use web server tool that automates the design of optimal PCR primers for several distinct USER cloning-based applications. Our web server, named AMUSER (Automated DNA Modifications with USER cloning), facilitates DNA assembly and introduction of virtually any type of site-directed mutagenesis by designing optimal PCR primers for the desired genetic changes. To demonstrate the utility, we designed primers for a simultaneous two-position site-directed mutagenesis of green fluorescent protein (GFP) to yellow fluorescent protein (YFP), which in a single step reaction resulted in a 94% cloning efficiency. AMUSER also supports degenerate nucleotide primers, single insert combinatorial assembly, and flexible parameters for PCR amplification.
Hans Jasper Genee, Mads Tvillinggaard Bonde, Frederik Otzen Bagger, Jakob Berg Jespersen, Morten O. A. Sommer, Rasmus Wernerson, and Lars Rønn Olsen
Software-supported USER cloning strategies for site-directed mutagenesis and DNA assembly
Rasmus Wernersson: firstname.lastname@example.org