RESUMEN

Fifteen related ligation-independent cloning vectors were constructed for high-throughput cloning and purification of proteins. The vectors encode a TEV protease site for removal of tags that facilitate pro tein purification (his-tag) or improve solubility (MBP, GST). Specialized vectors allow coexpression and copurification of interacting proteins, or in vivo removal of MBP by TVMV protease to improve screening and purification. All target genes and vectors are processed by the same protocols, which we describe here.

Asunto(s)

Vectores Genéticos , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Células/metabolismo , Clonación Molecular/métodos , Endopeptidasas/genética , Reacción en Cadena de la Polimerasa/métodos , Proteínas Recombinantes/biosíntesis , Transformación Genética