Genetic Transformation, TILLING (Targeting Induced Local Legions in Genomes) and VIGS (Viral Induced Gene Silencing) can all be used as reverse genetic tools to validate the function of pre-determined gene sequences in specific plant genotypes. Each method has specific strengths and weaknesses. They are also technologies that allow the conversion of basic research knowledge into crop species and potentially new commercial varieties. This document outlines the capacities and competencies of UK academic laboratories to apply these techniques for research (and in specific cases, commercial exploitation) and details the associated materials and resources available.

In addition, I have conducted a survey of the current status of cereal transformation in European Laboratories . This will facilitate access to collaborations and costed services for the transformation of cereals.

The most common use of genetic transformation in research is as a reverse genetic tool to assess the role of a defined gene sequence in any particular crop genotype. This can be done by over-expression of a transgene (which may be from the same or a different species), either constitutively or using tissue-specific, developmentally-regulated or inducible promoters. An alternative approach is to silence the expression of a specific native gene in a particular variety, again either constitutively or in a targeted manner. A refinement of these basic transgenic methods enables proteins encoded by transgenes to be targeted to specific sub-cellular compartments using transit peptides or to be tagged using immuno-reactive, short peptides with good, commercially-available antibodies.

The genetic transformation of plants, and of crop species in particular, is technically exacting and requires significant resources and organisational infrastructure. Services for the genetic transformation of several Monocot species, including wheat, barley, and the cereal model, Brachypodium are available through the Monogram network at one of the centres of excellence already established in the UK; RRes, JIC, NIAB, IBERS and/or SCRI. Indeed, the UK's national wheat transformation capability is unrivalled internationally. See also www.bract.org for information on transformation of cereal and Brassicae crops.

As an indication of costs, delivery of 25 transgenic plants from a single construct will be somewhere between £5,000 and £15000, depending on exact the nature of the requirement, and the species concerned. Most of the transformation centres have a long history of successfully delivering transgenic events to in-house and international collaborators and these services come with significant intellectual and material resources accrued through many years experience.

To further facilitate accessibility of transformation technologies, MONOGRAM Working Group IV aims to document and make available a wide range of transformation resources including: DNA constructs, promoter sequences, reporter genes and plant selectable markers, along with information about their application in monocots. Below are listed the species for which there is transformation capability in one or more UK institution and lists of resources available.

Viral induced gene silencing (VIGS) is a high-throughput method to transiently silence plant genes in susceptible tissues and can be used to study plant gene function. Virus-based vectors carrying sequences homologous to endogenous genes trigger silencing through a homology-dependent RNA degradation mechanism. The Barley stripe mosaic virus (BSMV), or less commonly, the Brome mosaic virus (BMV) may be used as a VIGS vector for cereals. The advantages of this transient knock-down approach are the rapidity of VIGS-induced symptoms (10d to 4weeks) and the capacity for relatively high-throughput studies.

VIGS has proved especially useful to study disease resistance genes in barley, and this technology has been used at SCRI. At RRes there is an intention to establish the VIGS system as a tool for reverse genetics studies in wheat, einkorn wheat (Triticum monococcum) and barley in a new facility, designed to operate at containment category 3 level, situated in the recently completed Jenkinson Building. This new facility comprises four walk-in controlled environment plant growth rooms and two laboratories (molecular biology and microscopy). Construction of this facility was jointly sponsored by the BBSRC and the Lawes Agricultural Trust. JIC and possibly IBERS are also interested in developing this area.

• VIGS at Rothamsted (RRes)

Targeted Induced Local Lesions IN Genomes (TILLING) is a non-transgenic method for inducing and identifying novel genetic variation. It is commonly used as a reverse genetic tool to study the role of known plant genes in specific genetic background. It has the unique advantage of allowing the generation of an allelic series of the target gene that potentially confer a range of phenotypes. The method has two distinct stages, firstly the generation of a TILLING library of DNA from a mutagenised population (or generated by natural variation, so-called EcoTILLING), and secondly, the identification of novel SNPs in the target genes using PCR-, HPLC-, temperature-, or resequencing-based approaches.