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Mutants are the classical and probably the most reliable genetic tool for assessing biological phenomena in living organisms. In tomato (Solanum lycopersicum L. Syn. Lycopersicon esculentum Mill.) the structure of the plant allows detection of a large array of hereditary variations. A plethora of mutant genes was thus identified and used for basic and applied studies from the early XX century. Work with mutants, however, tends to be limited by the difference in genetic backgrounds, which precludes reliable comparative studies. The various genetic backgrounds in tomato include hybrids, heirlooms and wild species. In 1998 we started to set up a collection of tomato mutants in a single genetic background, the cv Micro-Tom (MT).
MT is a miniature dwarf determinate tomato cultivar, released in 1989 by J. Scott and B. Harbaugh for home gardening purposes and launched as a genetic tool by the Avraham Levi group in 1997. On top of the relatively small tomato genome (950Mb), MT shares features with Arabidopsis that make it a suitable model system, such as small size (8 cm when grown in 50-100 mL pots) and short life cycle (70-90 days from sowing to fruit-ripening). These features allow the use of MT for large scale mutagenesis, since it is possible to grow a large population of MT lines using reduced spaces and to obtain advanced backcross generations in less time.
M1 plants growing in greenhouse at the density of 78 plants/m2. This density, which is considered to be low for MT, improves the number of M2 seeds harvest per individual M1 plants. Each bench has 460 adult (35-days-old) plants. Water supply only in the bottom of the pots prevents many fungal and bacterial diseases.
We used seeds treated with EMS (0.7%) and pollen treated with gamma-ray (80 Gy) to created M2 families (Pino-Nunes et al. Acta Horticulture, 821:63-72, 2009). Such families are being sown in the glasshouse or in the field for seedling or adult plant screening, respectively. Only those phenotypes observed in individual M2 plants that are confirmed in groups of derived M3 plants are included in our mutant collection.
Additionally, many well-characterized mutations previously discovered in other cultivars were introgressed (through at least six backcrosses) into MT. Our program of induced mutagenesis and introgression of known mutations is complemented by high throughput transgenic plant production (Pino et al. Plant Methods, 6:23, 2010) with emphasis in the expression of genes that alter hormone content/signaling and other genes governing plant development and responses to the environment.
Adult (80-days-old) M2 plants growing in field at Nunhems, Paulínina, SP Brazil. M3 seeds were collected from individual M2 plants presenting alterations in fruit traits (color, shape, ripening, brix).
Besides induced mutations, natural genetic variation is a valuable resource in tomato, especially for those loci where cultivated tomatoes already harbor knock-out (i.e. non-functional) versions of the genes. Such variation can be found in the tomato wild related species, most of which are inter-fertile and amenable to crossing with cultivated tomato. These species can be used to discover quantitative trait loci (QTL) and allelic variation for major genes. These Solanum (Sec. Lycopersicum) species evolved in South America where a large range of environments are present, representing thus a repository of genes related to different adaptation strategies. Using the same approach already developed to introgress mutations present in other cultivars, we are introgressing many monogenic natural genetic variation from the wild species S. galapagense, S. habrochaites, S. pennellii and S. pimpinellifolium into MT.
Tomato related wild species showing evident natural genetic variation in both vegetative (leaf shape) and reproductive (flower size and inflorescence architecture) development. Most of these variations are controlled by single locus where wild species harbor novel alleles.
As any other cultivated plant, the process of domestication of tomato replaced many “wild type” alleles to “mutated” ones. Thus, every tomato cultivar holds various mutant alleles when compared to any other genotype and this genetic assortment is exactly what defines a cultivar. In the case of MT, the best known mutant alleles already present in this cultivar are: dwarf (d), a brassinosteroid (BR)-related mutation responsible for the small plant size, and self-pruning (sp), responsible for its determinate growth habit.
There is no indicative for GA mutations controlling MT small size, since it has normal seed germination and leaf morphology. Additional alleles present in MT are uniform ripening (u), Stemphylium resistance (Sm) and Immunity to Fusarium wilt (I), which confer the absence of green shoulders in fruits and resistance to the pathogenic fungi Stemphylium solani and Fusarium oxysporum f. lycopersici, respectively. Some of these allelic variations are also present in other cultivars, as is the case of the the wide used cv M82, which harbors sp, u, I and Ve. Even for cultivars with no apparent mutations, such as Ailsa Craig and MoneyMaker, allelic variations exist at least in quantitative trait loci (QTL) controlling fruit size, since their fruits are larger than the fruits of the wild-type tomato (Solanum lycopersicum var. cerasiforme).
It is worth noting that with an appropriate control, the presence of mutations or allelic variations in a cultivar does not preclude it from being used to study the effects of specific mutations. However, if the event under study is influenced by that particular mutation, the alternative of generating near isogenic lines (NILs) harboring the non-mutated allele fulfills the requirement of an appropriate control in the scientific method. In the case of MT, NILs with indeterminate growth habit (MT-Sp), green-shoulder fruits (MT-U) and increased BR levels (MT-D) are available, among many others, in our mutant collection.
Micro-Tom lines harboring the wild type alleles D, Sp and U. The original MT line (bottom left and top right) holds the alleles d, sp and u, which confer dwarf plant size, determinate flowering and absence of green-shoulder fruits, respectively.
The late Charles Rick (1915-2002) once said that "if Arabidopsis is the Drosophila of plant genetics, then tomato has become the mouse". Given its small size and its precocity, the true equivalent of mouse for plant scientists is, in our opinion, Micro-Tom.