TransposonDs‐Mediated Insertional Mutagenesis in Rice(Oryza sativa)

Abstract

Rice (Oryza sativa) is the most important consumed staple food for a large and diverse population worldwide. Since databases of genomic sequences became available, functional genomics and genetic manipulations have been widely practiced in rice research communities. Insertional mutants are the most common genetic materials utilized to analyze gene function. To mutagenize rice genomes, we exploited the transpositional activity of an Activator/Dissociation (Ac/Ds) system in rice. To mobilize Ds in rice genomes, a maize Ac cDNA was expressed under the CaMV35S promoter, and a gene trap Ds was utilized to detect expression of host genes via the reporter gene GUS. Conventional transposon‐mediated gene‐tagging systems rely on genetic crossing and selection markers. Furthermore, the activities of transposases have to be monitored. By taking advantage of the fact that Ds becomes highly active during tissue culture, a plant regeneration system employing tissue culture was employed to generate a large Ds transposant population in rice. This system overcomes the requirement for markers and the monitoring of Ac activity. In the regenerated populations, more than 70% of the plant lines contained independent Ds insertions and 12% expressed GUS at seedling stages. This protocol describes the method for producing a Ds‐mediated insertional population via tissue culture regeneration systems. © 2016 by John Wiley & Sons, Inc.

Mutagenesis in Rice: The Basis for Breeding a New Super Plant

“…The transposon insertional activity has been widely applied to produce large scale mutations in rice genomes, as reviewed by Wang and collaborators (2013), which described that transposons such as miniature Ping ( mPing ), 607-pb and nDart1-3 were used for rice mutant line development. Also, two components of the maize transposon system, Activator / Dissociation ( Ac / Ds ) and Enhancer/Suppressor Mutator ( En/Spm-dSpm ) have been widely applied to produce insertional mutants in rice (reviewed in Wang et al, 2013; Xuan et al, 2016). In the presence of Ac transposase , the Ds element tends to be transposed and randomly integrated into the genome (McClintock, 1950).…”

“…In the presence of Ac transposase , the Ds element tends to be transposed and randomly integrated into the genome (McClintock, 1950). The Ac / Ds transposable system was applied in rice to produce mutagenesis (Upadhyaya et al, 2006; Xuan et al, 2016). In generated populations, more than 70% of mutant lines showed Ds -independent insertions.…”

The Functional Impact of Transposable Elements on the Diversity of Plant Genomes

“…Over the years, many such knock-out mutations were described, affecting a range of plant organs, including numerous reports on flower color and morphology [34][35][36][37][38]. The propensity for induction of knock-out mutations at a high rate made some TE families, for example, Ac/Ds and En/Spm from maize, routine tools for insertional mutagenesis, referred to as transposon tagging [39]. Robust collections of mutants were produced in different plant species including the model plant Arabidopsis thaliana (e.g., [40,41]).…”

Advances in omics technology for improving crop yield and stress resilience

“…Lines with new insertions but no active transposase are selected among the progeny of the cross. Different transposon systems as Activator/Dissociation ( Ac/Ds ), Suppressor‐mutator ( Spm ), Mu ( Mu ‐like elements), Tos retrotransposons, etc., are commonly used in several crop species like cereals, legumes and vegetables (Ayliffe & Pryor, 2011; Kishchenko et al, 2010; Mazier et al, 2007; Xuan et al, 2016; Zhu et al, 2007).…”