title: Positive-negative-selection-mediated gene targeting in rice.
abstract: Gene targeting (GT) refers to the designed modification of genomic sequence(s) through homologous recombination (HR). GT is a powerful tool both for the study of gene function and for molecular breeding. However, in transformation of higher plants, non-homologous end joining (NHEJ) occurs overwhelmingly in somatic cells, masking HR-mediated GT. Positive-negative selection (PNS) is an approach for finding HR-mediated GT events because it can eliminate NHEJ effectively by expression of a negative-selection marker gene. In rice-a major crop worldwide-reproducible PNS-mediated GT of endogenous genes has now been successfully achieved. The procedure is based on strong PNS using diphtheria toxin A-fragment as a negative marker, and has succeeded in the directed modification of several endogenous rice genes in various ways. In addition to gene knock-outs and knock-ins, a nucleotide substitution in a target gene was also achieved recently. This review presents a summary of the development of the rice PNS system, highlighting its advantages. Different types of gene modification and gene editing aimed at developing new plant breeding technology based on PNS are discussed.
authors.affiliation: Laboratory of Plant Molecular Genetics, Graduate School of Biological Sciences, Nara Institute of Science and Technology Ikoma, Japan.
authors.forename: Zenpei
authors.initials: Z
authors.lastname: Shimatani
authors.affiliation: Plant Genome Engineering Research Unit, Agrogenomics Research Center, National Institute of Agrobiological Sciences Tsukuba, Japan.
authors.forename: Ayako
authors.initials: A
authors.lastname: Nishizawa-Yokoi
authors.affiliation: Plant Genome Engineering Research Unit, Agrogenomics Research Center, National Institute of Agrobiological Sciences Tsukuba, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Endo
authors.affiliation: Plant Genome Engineering Research Unit, Agrogenomics Research Center, National Institute of Agrobiological Sciences Tsukuba, Japan.
authors.forename: Seiichi
authors.initials: S
authors.lastname: Toki
authors.affiliation: Development of Agrobiological Resources, Faculty of Agriculture, Meijo University Nagoya, Japan.
authors.forename: Rie
authors.initials: R
authors.lastname: Terada
keywords: gene editing; gene targeting; homologous recombination; knock-in; marker-free; positive–negative selection; rice; site specific recombination