title: Use of the computer-retargeted group II intron RmInt1 of Sinorhizobium meliloti for gene targeting.
abstract: Gene-targeting vectors derived from mobile group II introns capable of forming a ribonucleoprotein (RNP) complex containing excised intron lariat RNA and an intron-encoded protein (IEP) with reverse transcriptase (RT), maturase, and endonuclease (En) activities have been described. RmInt1 is an efficient mobile group II intron with an IEP lacking the En domain. We performed a comprehensive study of the rules governing RmInt1 target site recognition based on selection experiments with donor and recipient plasmid libraries, with randomization of the elements of the intron RNA involved in target recognition and the wild-type target site. The data obtained were used to develop a computer algorithm for identifying potential RmInt1 targets in any DNA sequence. Using this algorithm, we modified RmInt1 for the efficient recognition of DNA target sites at different locations in the Sinorhizobium meliloti chromosome. The retargeted RmInt1 integrated efficiently into the chromosome, regardless of the location of the target gene. Our results suggest that RmInt1 could be efficiently adapted for gene targeting.
authors.affiliation: Grupo de Ecología Genética, Estación Experimental del Zaidín; Consejo Superior de Investigaciones Científicas; Granada, Spain.
authors.forename: Fernando M
authors.initials: FM
authors.lastname: García-Rodríguez
authors.affiliation: Grupo de Ecología Genética, Estación Experimental del Zaidín; Consejo Superior de Investigaciones Científicas; Granada, Spain.
authors.forename: Teresa
authors.initials: T
authors.lastname: Hernández-Gutiérrez
authors.affiliation: Grupo de Ecología Genética, Estación Experimental del Zaidín; Consejo Superior de Investigaciones Científicas; Granada, Spain.
authors.forename: Vanessa
authors.initials: V
authors.lastname: Díaz-Prado
authors.affiliation: Grupo de Ecología Genética, Estación Experimental del Zaidín; Consejo Superior de Investigaciones Científicas; Granada, Spain.
authors.forename: Nicolás
authors.initials: N
authors.lastname: Toro
mesh_terms: D000465:Algorithms; D020029:Base Pairing; D001483:Base Sequence; D001665:Binding Sites; D002876:Chromosomes, Bacterial; D019295:Computational Biology / Q000379:methods*; D005091:Exons; D018390:Gene Targeting; D007438:Introns; D059372:Nucleotide Motifs; D056510:Position-Specific Scoring Matrices; D016962:Sinorhizobium meliloti / Q000235:genetics*
keywords: RmInt1; Sinorhizobium meliloti; gene targeting; group II introns; ribozyme