REVIEW OF LITERATURE

Transcription

1 REVIEW OF LITERATURE The Members of the Solanaceae family are in agriculture are tomato (Solanum lycopersicum and Solanum pimpinellifolium), pepper (Capsicum spp), eggplant (Solanum melongena), and potato (Solanum tuberosum); pharmacologically significant are tobacco (Nicotiana tabacum) and mandrake (Mandragora officinarum). Potato is the economically most important species within the Solanaceae. Potato is rich source of starch, protein, antioxidants, and vitamins. Tomato is second most consumed vegetable; it is important due to its dietary source of lycopene, beta-carotene, vitamin C and fiber. Solanum lycopersicum and S. pimpinellifolium (only two species with red fruits due to presence of lycopene) are also used as model species for fundamental research on disease resistance, pathogen response, stress tolerance, and fruit quality and development; wild S. pimpinellifolium is the closest wild progenitor of domesticated Solanaceae plants and together they are widely used in tomato breeding as a source material (Tanksley et al., 1992; Livingstone et al., 1999). The nuclear genome of potato and tomato consists of twelve chromosomes. Their genomes are measuring approximately 840 Mb and 950 Mb in size, respectively. Several large-scale rearrangements have been identified between the two genomes. Potato and tomato chromosomes display a similar morphology. The era of cost-effective next-generation sequencing technologies has enabled the rapid sequencing of a large number of plant genomes. Developing creation tools for structural and functional annotations are essential for comparative and evolutionary studies (Tanksley et al., 1992; Doganlar et al., 2002). 2.1 Tomato and Potato genomics The Solanum lycopersicum genomics is in an exciting phase of development following the recent sequencing of the potato and tomato genomes. Tomato was originated in South America, and was spread to rest of the world to become one of the most extensively used vegetable crops. Tomato is used for both basic and applied plant research. Several genetic and genomic resources were available for tomato before the inception of the tomato genome sequencing project. Large germplasm collections consisting of numerous accessions of landraces of tomato are a source of valuable disease resistance and other genes that had been exploited by breeders to develop modern cultivated tomato varieties (Majoros and Salzberg, 2004). Chapter II 5 Review of Literature

2 2.2 Genome analysis The Sol Genomics Network (SGN; is a comparative genomics platform, with genetic, genomic and phenotypic information of the Solanaceae family and its closely related species that incorporates a community-based gene and phenotype curation system. Well-annotated multigene families are useful for further exploration of genome organization and gene evolution across species. As an example, the multigene transcription factor families, WRKY and Small Auxin Up-regulated RNA (SAUR), both play important roles in responding to abiotic stresses in plants (Gascuel et al., 2001; Gremme Genome Annotation et al., 2005; Database URL: The linear strings of nucleotides that together comprise a genome sequence are of limited interest by themselves. Genome annotation encompasses the process of assigning a plausible biological interpretation to a genome sequence through identification and characterization of the elements contained therein. Ideally, each element in a genome such as a gene or a transposon is annotated through experimentally obtained evidence. The disparity in throughput with which genomes can be sequenced and assembled compared to the laboriousness of experiments to determine the precise structure and function of a single element has resulted in the development of algorithms that can predict these features in a genome (Majoros and Salzberg, 2004; Korf, 2004; Lomsadze et al., 2005; Stanke et al., 2008). Genome annotation include structure annotation and function annotation where structural genome annotation is the determination of the precise position, boundaries and composition of different sequence elements, whereas function annotation attempts to assign a probable biological function to each of these sequence elements. The quality of the WGS assembly through alignment to Sanger-derived phases 2 BAC sequences. In an alignment length of ~1 Mb (99.4% coverage), no gross assembly errors were detected. Alignment of cosmid and BAC paired-end sequences to the WGS scaffolds revealed limited ( 0.12%) potential misassemblies. Extensive coverage of the potato genome in this assembly was confirmed using available expressed sequence tag (EST) data; 97.1% of 181,558 available Sanger-sequenced S. tuberosum ESTs (>200 bp) were detected. Repetitive sequences account for atleast 62.2% of the assembled genome (452.5 Mb) with long terminal repeat retrotransposons comprising the majority of the transposable element classes, representing 29.4% Chapter II 6 Review of Literature

3 of the genome. In addition, subtelomeric repeats were identified at or near chromosomal ends (Figure 2.1) (The Potato Genome Sequencing Consortium, 2011). The genome of the inbred tomato cultivar Heinz 1706 was sequenced and assembled using a combination of Sanger and next generation technologies. The predicted genome size is approximately 900 megabases (Mb), consistent with previous estimates, of which 760 Mb were assembled in 91 scaffolds aligned to the 12 tomato chromosomes, with most gaps restricted to pericentromeric regions (Figure 2.2). Base accuracy is approximately one substitution error per 29.4 kilobases (kb) and one indel error per 6.4 kb. The scaffolds were linked with two bacterial artificial chromosomes (BAC)-based physical maps and anchored/oriented using a high-density genetic map, introgression line mapping and BAC fluorescence in situ hybridization (FISH) (The Tomato Genome Consortium, 2012). Annotation of a genome sequence involves the execution of a number of different tools in a particular combination and order on a collection of sequences, ranging from a small number of complete chromosomes to hundreds or thousands of sequence contigs and scaffolds. Genome sequences from evolutionary related species can readily be annotated using the same software tools albeit sometimes with modified parameters and reference data. These properties make genome annotation a repetitive, modular task with many inter-task dependencies that can be described (Burge and Karlin, 1997; Allen and Salzberg, 2005; Picardi and Pesole, 2010). Figure 2.1: The potato genome organization (Source: The Potato Genome Sequencing Consortium, 2011). Chapter II 7 Review of Literature

4 Figure 2.2. Tomato genome topography and synteny (Source: The Tomato Genome Consortium, 2012) 2.4 Structural and functional genome annotation All the features in a genome, protein-coding genes are the most extensively studied. In eukaryotes, the coding region may be interrupted by introns, which can be identified computationally through conserved signals on the border between the coding exons and non-coding introns (splice sites) (Warburton et al., 2004; Kofler et al., 2007). Once the elements in a genome sequence have been identified, the next step is to assign to them with a plausible biological function. Computational inference of the function of a particular sequence can be achieved either directly through sequence similarity searches, or indirectly through the identification of common motifs or domains between groups of functionally related sequences. Both methodologies exploit the wealth of sequence annotations that have been generated and deposited in public databases in the past decades to annotate a newly generated sequence. The accuracy and reliability of annotations derived from database searches depend Chapter II 8 Review of Literature

5 strongly on both the availability of evolutionarily related sequences in the databases, and the quality of their annotations. BLAST searches and can be used on many public as well as private sequence databases (Altschul et al.,1997; Benson et al., 2011; Magrane and Consortium, 2011). Motif and domain searches provide a more coarse-grained alternative to sequence similarity searches. While the latter focus on the similarity between two sequences over their whole length, the former rely on the conservation of small subsequences within a group of functionally related sequences. Prime examples of such conserved subsequences are protein domains, the modular functional sub-parts of proteins. Domains can be identified and extracted from a multiple sequence alignment of functionally related proteins and represented as HMMs or WMMs, which in turn can be used to query novel protein sequences (Hunter et al., 2009; Gene Ontology Consortium, 2010). 2.5 Comparative genomics of Solanum lycopersicum analysis The integration and advancements of molecular biology, evolution, and computer science over the past few decades have led to the development of several new fields of study. Comparative genomics, the study of the similarities and differences between two or more genomes, continues to be fueled by the rapidly growing number of fully sequenced genomes in databases. Comparative genomics has also been aided in the rapid advancements in sequencing technologies over the last few decades. Next Generation (NG) sequencing technologies have become a great resource to the genomics community because of the extremely low 'per base' cost of sequencing (Fickett and Tung, 1992; Guigo, 1998; Rice et al., 2000). In earlier studies on sequence analysis of Solanaceae gene families, P450 mono-oxygenases and serine threonine protein kinases were found to be overrepresented in potato as compared with tomato, and in both plants, the P450 genes were expanded much more than in Arabidopsis thaliana. Confirmation of computationally identified gene families with experimental data adds the next layer of annotation, and these genes can serve as significant anchors in the genome sequence for researchers looking for unknown genes located near the experimentally validated genes. Along with micro-synteny, the conserved order of genes and gene families across organisms are important data types for genome comparisons. Compared to the potato genome, the tomato and S. pimpinellifolium genomes showed more than 8% nucleotide divergence. Chapter II 9 Review of Literature

6 Moreover, the tomato genome is highly syntenic with the genomes of other economically important members of the family Solanaceae, such as eggplant and pepper. Comparative genome analysis identified two consecutive triplication events in the Solanum lineage. Interestingly, these genome triplications added new gene family members such as transcription factors and enzymes necessary for ethylene biosynthesis and perception, which mediate important fruit-specific functions (Eddy, 2001). The most common repeat families in the tomato libraries were the Gypsy ( %) and Copia ( %) classes of retrotransposons. Another prominent class of repeats comprised the ribosomal RNA genes (< %). The tomato Eco (EcoRI) library had the lowest repeat density at 13.0%, which can be attributed to a lower amount of Gypsy retrotransposons (5.0%). The highest repeat content was found in the tomato Mbo (MboI) library (22.9%), more than a third of which (8.6%) consisted of ribosomal RNA genes (Datema, 2011). An initial effort was made to compare the gene and repeat content of the tomato and potato genomes, based on the available BAC-end sequences for both species. The BAC-end sequence comparison is of particular interest as it provides a picture for the complete genome, including both euchromatic and heterochromatic sequence. Comparison using only sequenced tomato BACs will mainly provide a comparison between the euchromatin of tomato and potato. In total, 310,580 BAC-end sequences representing ~19% of the 950-Mb tomato genome were compared to 128,819 potato BAC end sequences representing ~10% of the 840-Mb potato genome (Song et al., 2000; Chen et al., 2004). It is important to note that while most potato varieties used in agriculture are tetraploid, the potato line being sequenced is diploid. The tomato genome has a higher overall dispersed repeat content than the potato genome, with the majority of dispersed repeats in both species belonging to the Gypsy and Copia retrotransposon families. On the other hand, simple sequence repeats (SSRs) motifs are more abundant in potato than in tomato. In both genomes, penta-nucleotide repeats are the most common form of SSRs, and AAAAT is the predominant repeat motif. This is in contrast to previously studied plant species, in which di- and penta-nucleotide repeats generally occur least frequently. Taking into account the difference in genome size and assuming that tomato has ~40,000 genes, potato appears to contain up to 6400 more putative coding regions than tomato. Moreover, the P450 superfamily appears to have expanded dramatically in both species compared with Chapter II 10 Review of Literature

7 Arabidopsis thaliana, suggesting an expanded network of specialized metabolic pathways in the Solanaceae (Kanyuka et al., 1999; Song et al., 2000; Chen et al., 2004). The recently published genome sequences of tomato and potato have shed new light on chromosomal organization in the Solanaceae family, and provide insights into the evolution of plant genomes. 2.6 Genome sequencing and annotation Potato is the first sequenced genome of an Asterid, a clade within Eudicots that encompasses nearly 70,000 species characterised by unique morphological, developmental and compositional features. In potato genome the total number of 39,031 protein-coding genes were predicted (The Potato Genome Sequencing Consortium, 2011) while in tomato genome the total number of protein-coding genes is 34,727 (The Tomato Genome Consortium, 2012) and identified 18,320 orthologous tomato potato gene pairs. 2.7 Improvement for plant resistance against abiotic stresses Plant response to abiotic stress is the result of complex synchronized actions of gene networks. Drought is a worldwide problem and understanding the processes underlying drought stress tolerance in plants is a high priority in many research projects. Drought resistance is a complex process, and little is known about the molecular mechanisms underlying the plant response and tolerance. The responses involve biochemical, physiological, molecular, cellular and whole-plant changes. Genetic, molecular and genomic analyses of drought response and tolerance in a number of plants such as Arabidopsis, rice, maize, tomato and other plants have revealed several drought-inducible genes that appear to play different roles in managing drought stress (Duque et al., 2013). 2.8 Basic facts about the Potato Genome Cultivated potato has a chromosome number of 2n = 4x = 48, and a haploid genome size of 850Mb, roughly six times that of Arabidopsis thaliana and twice the size of the rice genome. The potato genome is very similar in size to its close relative tomato, and genetic maps of the two species show high levels of macrocolinearity. Two genomes are conserved at the microsyntenic level should start to become available as outputs from the respective genome projects accumulate. The tomato genome mainly comprises low-copy-number sequences, which diverged rapidly in evolutionary time (Wang et al., 2005; Zhu et al., 2008). Chapter II 11 Review of Literature

8 It is also known that the majority of tomato heterochromatin is found in centromeric regions with almost all of the euchromatic DNA located distally in long uninterrupted tracts, a structural feature likely to be true of potato. Gene isolation and recent BAC-end sequencing efforts are providing the first detailed glimpses of the genome structure in potato. Using BAC-end sequence and full BAC sequence data, it has also been shown that potato (34%) contains considerably less repetitive DNA than tomato (46%), this difference being consistent with relative genome sizes of the two crops (850 versus 1000Mb, resp.) (Tanksley et al., 1992; Gavrilenko, 2007). Schweizer et al. (2009) who characterised the potato genome in terms of the amounts of different classes of repetitive DNA, suggest that the more highly repeated sequences comprise only 4 7% of the potato genome, suggesting that it was relatively devoid of highly repetitive DNA sequences, thus supporting the earlier tomato study. The generation of large expressed sequence tag (EST) collections is a primary route for large-scale gene discovery. There have been several efforts to generate EST resources for potato. The potato gene index ( compbio.dfci.harvard.edu/tgi/cgi-bin/tgi/gimain.pl?) contains almost ESTs, assembled into more than contigs with over singletons. These efforts, while not exhaustive, comprise a major genomics resource for potato researchers, perhaps comprising between 50 70% of the total potato gene repertoire. These ESTs will form an important source not only for the discovery of candidate genes and genetic markers, but also for the development of microarrays, until the whole genome sequence becomes available in potato. The tomato and potato sequencing projects will have huge implications for those working in the Solanaceae, and will further sharpen the requirement for functional genomics tools (Van der Hoeven et al., 2002). Potato, a highly heterozygous tetraploid, is undergoing an exciting phase of genomics resource development. The potato research community has established extensive genomic resources, such as large expressed sequence tag (EST) data collections, microarrays and other expression profiling platforms, and large-insert genomic libraries. Moreover, potato will now benefit from a global potato physical mapping effort, which is serving as the underlying resource for a full potato genome sequencing project, now well underway. These tools and resources are having a major impact on potato breeding and genetics. The genome sequence will provide an invaluable comparative genomics resource for cross-referencing to the other Solanaceae, notably tomato, Chapter II 12 Review of Literature

9 whose sequence is also being determined. Most importantly perhaps, a potato genome sequence will pave the way for the functional analysis of the large numbers of potato genes that await discovery. Potato, being easily transformable, is highly amenable to the investigation of gene function by biotechnological approaches (Zamir and Tanksley, 1988; Schweizer et al., 1993). The available potato EST resources comprise an unknown but significant fraction of the gene complement of potato, and are derived from several genotypes, tissues, and environmental influences. A nonredundant set of of these ESTs was used by the Institute for Genomic Research (TIGR) to develop a cdna potato microarray that was made available to the research community at minimal cost. Moreover, the same organisation offered a transcription profiling service to allow the evaluation of these arrays by a wide range of users working on different Solanaceous plant species asking different biological questions. This allowed generation of massive microarray data that is publicly available ( service2.shtml#aprocedure). These studies, while informative, highlight the dilemma faced by plant molecular biologists in prioritizing genes for further study from a large number of candidate genes in the absence of genetic information and mutations in target trait genes (Arumuganathan and Earle, 1991). 2.9 Functional Studies in Potato Potato geneticists and breeders have generated a great deal of information about the location of genes and QTLs coding for important potato traits, including pest and disease resistance and tuber traits. Developments in genetics and structural genomics are beginning to be matched by concomitant development of functional genomics tools. Potato has a strong need for a highdensity gene map or a genome sequence, to place gene sequences in their genetic/genomic context. Relatively high-throughput methods are also needed for testing and assessing gene function. The availability of mutant populations of potato will also be of tremendous value in this regard (Simko et al., 2006; Muth et al., 2008). Potato cultivars are highly heterozygous and contain very high levels of genetic load. It has been estimated that there is one SNP approximately at every 25 bp. There have been some recent tantalizing developments in functional genetics/genomics tools and resources for potato. Gene expression profiling or microarray studies have a role to play in the identification of a Chapter II 13 Review of Literature

10 pool of candidate genes potentially involved in any given biological process. These methods, in combination with other functional genomics tools such as RNA interference (RNAi), virus-induced gene silencing (VIGS), and activation tagged lines, have the potential to facilitate the identification of the role of thousands of potato genes over the next several years (Simko et al., 2006; Muth et al., 2008). Potato has entered an exciting new era, whereby the development of extensive genetic and genomic resources have opened up many new possibilities for studying important potato traits relevant to potato agronomy. Concomitant development of similar resources for other Solanaceous species, notably tomato, and a growing cohesiveness of the Solanaceae research community, as demonstrated by the SOL vision bode well for future genomic research of potato and its close relatives ( Development of biotechnological tools for assaying potato gene function is likely to progress rapidly in the coming years (Simko et al., 2006; Muth et al., 2008) Computational approaches for plant breeding and genome research The greatest challenge facing the molecular biology community today is to make sense of the wealth of data that has been produced by the genome sequencing projects. Traditionally, molecular biology research was carried out entirely at the experimental laboratory bench but the huge increase in the scale of data being produced in this genomic era has realized a need to incorporate computers into the research process. With the advent of new tools and databases in molecular biology we are now enable to carry out the research not only at genome level but also at proteome, transcriptome and metabalome levels. Bioinformatics is an interdisciplinary area of the science composed of biology, mathematics and computer science. Bioinformatics is the application of information technology to manage biological data that helps in decoding plant genomes. During the last decades enormous data has been generated in biological science, firstly, with the onset of sequencing the genomes of model organisms and secondly, rapid application of high throughput experimental techniques in laboratory research. Biological research that earlier used to start in laboratories, fields and plant clinics is now starts at the computational level using computers for analysis of the data, experiment planning and hypothesis development. Application of various bioinformatics tools in biological research enables storage, retrieval, analysis, annotation and visualization of results and promotes better understanding of biological Chapter II 14 Review of Literature

11 system in fullness. This will help in plant health care based disease diagnosis to improve the quality of Plant life. The sequencing of the genomes of plants and animals will provide enormous benefits for the agricultural community. Bioinformatics tools can be used to search for the genes within those genomes that are useful for the agricultural community and to elucidate their functions. This specific genetic knowledge could then be used to produce stronger, more drought, disease and insect resistant crops and improve the quality, making them healthier, more disease resistant and more productive (Martienssen, 2004). Bioinformatics is the key for realizing the full potential of post-genomic revolution moving plant science toward crop systems biology. It will help in exploring the benefit of bioinformatics application to plant research and, particularly, to crop science. Plant biologists and information technology specialists can contribute equally to such a task by organizing their work in a collaborative and interdisciplinary manner, thus applying in the most effective way their different technical skills to solve agricultural problems. Genomics, proteomics, metabolomics can be applied for data integration known as phenome that comprises all the layers of the phenotype which is studied as a whole and arise from the interaction of the genome with the environment. As for all the other omics, the phenome of a plant thus represents the sum total of its phenotypic traits (Faccioli et al., 2009). InterProScan employs a large collection of protein domain databases to identify conserved protein signatures in sequences of interest. Software like MUMmer and BLASTZ/LASTZ has been developed to align complete genome sequences and extract the variation between them. Tools to predict gene sequences in a genome range from naïve Open Reading Frame (ORF) predictors like getorf from the EMBOSS suite to complex eukaryotic gene finders such as geneid, genscan and Glimmer HMM ( Comparisons of gene content and gene order Gene content comparisons were performed with Multipipmaker (Schwartz et al., 2003), GenomeThreader, EuGene and JIGSAW is used for identification and masking of repetitive elements in a genome sequence a database of known repeat sequences, for example using the RepeatMasker software with the RepBase database. The RECON software identifies repeats through their multiplicity in the genome sequence, without taking into account a prior knowledge Chapter II 15 Review of Literature

12 about the structure of the elements. Annotation of the potato and tomato chloroplast genomes was performed using DOGMA (Dual Organellar GenoMe Annotator; jgipsf.org/dogma). An aligned data set of all of the shared genes among the four Solanaceae chloroplast genomes was constructed by extracting these sequences from the annotated genomes either using DOGMA or the Chloroplast Genome Database. The sequences were aligned using ClustalX followed by manual adjustments using Seq Ap Comparison of intergenic spacer regions Intergenic regions from four Solanaceae chloroplast genomes were compared using MultiPipMaker ( tools.html). The modern biologist works in an unprecedented time in the history of scientific knowledge. There are several factors that have been converging over the last few decades that have increased scientific productivity in a profound way. First, molecular biology has continued to be advanced through the tools and techniques that help researchers to study genetics at the molecular level. Finally, the large volume of sequence data combined with the continued advances in the desktop computer, have led to the integration of molecular biology, evolution, and computer science. This integration has led to the development of several new fields of study including bioinformatics, genomics, and comparative genomics. Bioinformatics is the use of computer science, mathematics, and information theory to model and analyze biological systems. While genomics is the study of an organism s entire genome, comparative genomics is the study of similarities and differences between two or more genomes. PlantTribes offers a unique view of gene families and plant genomes that facilitate comparative analyses. Additional information is connected to each sequence; including domain presence from NCBI s Conserved Domain Database (CDD). The PlantTribes database offers a unique and powerful view of plant genomes and evolution. The automated annotation of the genomes provide an important source of information for gene analysis; however, automated methods do not yet produce perfect results and sometimes require intervention using manual curation. Tools used for this purpose include GenomeView and Apollo ( tools.html). Chapter II 16 Review of Literature

13 2.13 Plant genomes : current status Plant breeding and genetics are powerful tools for increasing plant productivity through development of improved varieties. The rapid progress of plant genomics in recent years has opened new possibilities in targeted breeding of specific traits, and provides a powerful approach to sustainable crop production. Plant genomics, in combination with genetics and breeding, has a particularly crucial role to play in ensuring food security to the rapidly growing world population. Many plant genomes are large and complex due to an abundance of transposable elements and a long history of repeated genome duplication, making genome sequencing a major challenge. The era of plant genomics began with release of the Arabidopsis genome sequence in It was a milestone in plant biology and made Arabidopsis one of the most popular species for basic plant research. Rice, a staple food in most of the world, was the second available plant genome in However, members of the group Euasterids, which has many plants of economic importance, were not represented in the list of known plant genome sequences until the release of the potato (Solanum tuberosum) genome belonging to the family Solanaceae. The recently decoded genome sequences of tomato (Solanum lycopersicum) and its close wild relative (Solanum pimpinellifolium) are significant additions to published Euasterid genomes. These genomes will not only promote plant genomics and breeding studies for crop improvement programs in the Euasterids, particularly in the family Solanaceae, but also provide an unprecedented opportunity for basic plant biological research in the area of development and evolution ( Future directions of tomato genetics and genomics Modern tomato genetics had already used molecular markers and functional analysis to identify a handful of genes underlying developmental or yield traits, but the availability of the tomato genome sequence will further revolutionize tomato genetics and breeding. This will not only help to identify useful SNPs from the wild accessions but also identify rare SNPs within domesticated varieties. Tomato breeders can then target gene variants (SNPs) in the wild species associated with desirable traits such as disease or pest resistance or growth in extreme environmental conditions and introduce them into cultivars in order to exploit the rich tomato germplasm for breeding purposes. More genome sequences will facilitate QTL identification, mapping and cloning of underlying genes, and provide new SNP markers for Chapter II 17 Review of Literature

14 marker-assisted breeding. Additionally, millions of informative markers (SNPs/InDels) and structural variations, such as duplications, inversions, transpositions, and so on, identified through comparison of genome sequences of domesticated and wild tomatoes will promote investigations into the genetic and molecular basis of the process of domestication and crop improvement. Integrating additional functional genomics approaches such as metabolomics and proteomics can significantly reduce the number of candidate genes for a given QTL. One of the major thrusts of functional genomics in future will be RNA-seq enabled transcriptome profiling. For example, comparison of transcriptome profiles from domesticated and wild tomato species will give us insights into the gene expression differences associated with the process of domestication and trait diversity. The tomato functional genomics database (TFGD), which includes microarray, metabolite and small RNA data, has already been established as a comprehensive resource even before the complete tomato genome sequence was released. Availability of the tomato genome sequence will speed up the understanding of gene function in developmental and metabolic pathways and identify key steps in co-regulation mechanisms by mapping relevant tomato mutants. Additionally, multiple TILLING (Targeting Induced Local Lesions IN Genomes) resources in different backgrounds have already been developed for tomato functional genomics. These TILLING resources, in combination with the tomato genome sequence, should be useful for both forward and reverse genetics in tomato for both basic science and/or crop improvement (Aashish et al., 2012). Chapter II 18 Review of Literature