Jean-Michel Ane´ et al. with conserved cysteine motifs whose function continues to be investigated [111–113]. But as genome scale biology is applied, the need to syn- thesize transcriptomics data, proteomics data, metabolomics data, and more becomes as important as the availability of informatics tools to analyze these data individually. Several steps in this direction have occurred within the M. trun- catula community. Some of these integrated solutions are focused around a process, such as gene expression in my- corrhizal symbiosis [114]. Because of the nearly complete genome sequence and the cooperative nature of the sequenc- ing and annotation of the genome, comprehensive integra- tion of various data sources has been necessary from the be- ginning. Cannon et al. [115] provide a nice summary of the available sequence-based resources and how they interact. A freely available database of biochemical pathway data for M. truncatula (MedicCyc) contains more than 250 pathways with related genes, enzymes, and metabolites [116]. This pro- vides the ability to not only visualize metabolomics data and integrate them with functional genomics data, but also al- low comparison of M. truncatula pathways to those in other plants using the compatible AraCyc and RiceCyc databases. 5. REVERSE GENETICS Reverse genetics approaches which identify mutants in a gene of interest based on sequence diﬀerences are critical genomic tools in a model system. A range of approaches are available, including retrotransposon tagging, T-DNA tagging, TILL- ING for EMS mutations, PCR screening for fast neutron mu- tations, and RNA-induced gene silencing (RNAi) [46]. Each method has advantages and disadvantages, and the choice of which method(s) to use will depend on the purpose of the investigator. In M. truncatula, RNAi, TILLING, and PCR screening of Tnt1 insertion mutagenesis populations or fast- neutron generated deletion populations are reverse genetic approaches presently possible. As noted above, the eﬃcacy of RNAi in M. truncat- ula has been documented [48] including use in whole plants and in transformed roots. The combination of RNAi constructs and hairy root transformation is useful for large scale screening projects to identify genes of in- terest for further analysis. A large-scale project to iden- tify gene function by silencing in M. truncatula is under- way (http://www.cbs.umn.edu/labs/ganttlab/rnai.html). Ini- tial results from this project include identiﬁcation of a calcium-dependent protein kinase involved in nodule devel- opment, a gene that had not been identiﬁed through classical mutational analysis [117]. TILLING (Targeting Induced Local Lesions in Genomes) has proven useful in Arabidopsis and other plants (reviewed in [118]). Brieﬂy, the sequence of a gene of interest is an- alyzed with a computer program that determines the con- sequences of all possible EMS mutations (primarily G to A transitions) on the amino acid sequence of the deduced pro- tein. Regions are chosen for PCR ampliﬁcation based on the concept that those regions most likely to result in altered pro- tein function are highly conserved domains in proteins, and PCR primers are designed to amplify these regions of DNA 5 from each plant. The PCR products are analyzed for single- base pair changes in a high throughput sequencing gel system using an enzyme that detects and cleaves single-base mis- matches in DNA. The use of high throughput methodology and a well-characterized and curated population of mutag- enized plants allows a plant containing a lesion in the gene of interest to be identiﬁed in days. The beneﬁts of TILLING are not only the rapid identiﬁcation of lesions, but the nature of the lesions themselves. The point mutations generated by EMS treatment allow the use of TILLING to generate an al- lelic series that includes both missense and nonsense muta- tions. In M. truncatula, the Cook lab. at UC Davis developed a population of ∼4000 curated EMS mutagenized plants for purpose of TILLING. This resource is currently unavailable as a community resource due to the absence of funding. To date genotypic screens for mutations in 15 genes of inter- est to the Cook lab or collaborators have been undertaken, and 143 mutants identiﬁed, with recovery rates of 9.89 al- leles per kbp screened. Phenotypic characterization of one (of 23 unique) allele identiﬁed from one of the early geno- typic screens for mutations in the M. truncatula arbuscule speciﬁc phosphate transporter MtPT4 is described by Javot et al. [119]. Characterization of other mutants in this col- lection is currently ongoing or advanced to the stage where manuscripts are in preparation for submission (Douglas R. Cook and Varma Penmetsa, personal communication). A reverse-genetics platform has been established in Med- icago truncatula exploiting fast neutron (FN) mutagenesis and a highly sensitive PCR-based detection ﬁrst documented in Arabidopsis [120]. The FN-based screening platform pro- duces complete loss of function mutants by identifying large deletions in the targeted region. Central to this platform is the development of a detection strategy which allows a mu- tant amplicon, possessing an internal deletion, to be prefer- entially ampliﬁed in pools where genomic target sequence is present at a 20 000-fold excess. This detection sensitivity has been achieved through a combination of techniques for suppressing the ampliﬁcation of the wild-type sequence and preferentially amplifying the mutant product. The popula- tion has been arrayed such that 12 000 M2 plants can be an- alyzed in 4 PCR reactions. These megapools can then be dis- sected using 25 PCR reactions on 3D pools, allowing iden- tiﬁcation of the individual seed lot containing the mutant. In comparison with the well-established TILLING method [121–123], which utilizes 8-fold PCR-based screening, FN alleles can be isolated at a fraction of the cost and avoid the problems associated with EMS mutagenesis of target- ing small genes and the very high number of background mutations in isolated mutants. An initial characterization of the FN system analyzed 10 genes in a subpopulation of 60 000 M2 plants. Mutants were recovered for 4 target genes. A population of 180 000 M2 plants has now been established and should allow the recovery of mutants from a majority of targeted loci. Information for accessing this resource can be found at www.jicgenomelab.co.uk. (C. Rodgers and G. E. D. Oldroyd, personal communication). Recently, researchers have identiﬁed a tobacco retrotrans- poson, Tnt1, that moves randomly in M. truncatula but only