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Targeting Induced Local Lesions IN Genomes

TILLING (Targeting Induced Local Lesions IN Genomes) is a general reverse genetic technique that uses traditional chemical mutagenesis methods to create libraries of mutagenized individuals that are later subjected to high throughput screens for the discovery of mutations. We are based at the UC Davis Genome Center and will provide a TILLING service for japonica rice starting late Summer 2008. Hexaploid and tetraploid wheat will be added in the future. Tomato is in the works.

Placing a TILLING order

Click on the How to Place an Order link to go to our ordering page.

Background

The explosion of DNA sequence information has created a unique opportunity to investigate the function of genes. The approach that determines the function of genes first defined by DNA sequence analysis, is called reverse genetics. The tools available for reverse genetics of plants include the use of transposons or T-DNA for gene tagging and the use of RNA interference. While powerful, these methods have limitations. For example, they do not work in all plant species. Targeting Induced Local Lesions IN Genomes (TILLING) is reliable and widely applicable. TILLING combines chemical mutagenesis with mutation screens of pooled PCR products, resulting in the isolation of missense and nonsense mutant alleles of the targeted genes. TILLING has two significant advantages over existing plant gene knock-out tools: first, it is applicable to any plant since it does not require transgenic or cell culture manipulations. Second, it produces an allelic series of mutations including hypomorphic alleles that are useful for genetic analysis. The TILLING technology can be used to discover and survey natural variation. The method is called Ecotilling and has been applied to Arabidopsis and human (see Publications link in the side bar).

History

TILLING was originally conceived by Claire McCallum, a UW graduate student who wanted targeted mutations in the chromomethylase genes of Arabidopsis. Chuck Dearolf and collaborators independently conceived a similar project for Drosophila. Development of TILLING in Seattle was in collaboration with the Henikoff lab and this lead to the Seattle TILLING Project (STP), a FHCRC-UW collaboration that provided Arabidopsis and fruit flies mutants. With the departure of Luca Comai from Seattle (Summer 2006), STP is now operated by Steve Henikoff. The TILLING objectives have been split: diploid Arabidopsis and fruit flies remain at STP. The rice project has been moved to Davis.

Overview of TILLING at the UCD Genome Center

Public service TILLING for rice is now at the beta testing stage at the UC Davis Genome Center. Rice TILLING at UCD is carried our in collaboration with the laboratory of Tom Tai, USDA-ARS at Davis. Rice_Tilling was funded by USDA-NRI and the Rockefeller Foundation and is presently sustained on user's fees. Wheat TILLING has been developed at UCD in collaboration with Jorge Dubcovsky at the UCD department of Plant Sciences and it is funded by a USDA-NRI award. A service for wheat is being planned. Tomato _Tilling is in collaboration with Roger Chetelat and Allan VanDeynze at UC Davis and is funded by a USDA-NRI plant genome award to provide a service to the tomato community. Tetraploid Arabidpsis TILLING provides a novel approach to isolate KO mutants in genes that are required in the gametophyte and has been developed in collaboration with the labs Sundaresan Venkatesan (Sundar) and Simon Chan. Starting in the Fall of 2007, we have converted to TILLING by Sequencing and essentially abandoned the LiCor-CELI mismatch detection method. Research to develop the TILLING-by-Sequencing method using rice, wheat, and arabidopsis as a model system is now funded by an award from the NSF Plant Genome Program.

TILLING workshops

The TILLING-by-Sequencing project offers Training Workshops. The first was held Aug 24 and 25 2009 in Davis and had 9 participants. Additional ones are planned.

Funding

Funding for Rice and Tomato TILLING and the forthcoming service is by USDA CSREES, NRI Plant Genome Program and in part by a grant from the Rockefeller Foundation. TILLING-by-sequencing is funded by NSF.

Cooperative State Research, Education, and Extension Service