Updating the Micro-Tom TILLING platform

Abstract

The dwarf tomato variety Micro-Tom is regarded as a model system for functional genomics studies in tomato. Various tomato genomic tools in the genetic background of Micro-Tom have been established, such as mutant collections, genome information and a metabolomic database. Recent advances in tomato genome sequencing have brought about a significant need for reverse genetics tools that are accessible to the larger community, because a great number of gene sequences have become available from public databases. To meet the requests from the tomato research community, we have developed the Micro-Tom Targeting-Induced Local Lesions IN Genomes (TILLING) platform, which is comprised of more than 5000 EMS-mutagenized lines. The platform serves as a reverse genetics tool for efficiently identifying mutant alleles in parallel with the development of Micro-Tom mutant collections. The combination of Micro-Tom mutant libraries and the TILLING approach enables researchers to accelerate the isolation of desirable mutants for unraveling gene function or breeding. To upgrade the genomic tool of Micro-Tom, the development of a new mutagenized population is underway. In this paper, the current status of the Micro-Tom TILLING platform and its future prospects are described.

Keywords: Micro-Tom; TILLING; reverse genetics; tomato.

Fig. 1

Flow chart for constructing mutant populations with double-EMS mutagenesis. The conventional process of EMS mutagenesis (upper panel) was described by Saito et al. (2011). In the following steps, bulked M₃ (M₃M₀) seeds were mutagenized with 1.0% EMS and M₃M₁ seeds were sown in the greenhouse to harvest M₄M₂ seeds from single M₃M₁ plants. Ten M₄M₂ seeds were sown and grown as an M₄M₂ line and M₅M₃ seeds were harvested from the same M₄M₂ lines as a bulked seed. Mutant DNA was extracted from each M₄M₂ line (10 plants per line) and pooled as 8-fold DNA for TILLING screening.