Abstract
Resistance to begomoviruses, including bipartite tomato mottle virus (ToMoV) and monopartite tomato yellow leaf curl virus (TYLCV), has been introgressed to cultivated tomato (Solanum lycopersicum) from Solanum chilense accessions LA1932 and LA2779. A major gene, Ty-3, responsible for resistance to ToMoV and TYLCV was previously mapped on the long arm of chromosome 6. In the present study, we identified a 14-cM S. chilense introgression on the long arm of chromosome 3 in some resistant breeding lines derived from LA1932. A new begomovirus resistance locus, Ty-4, was mapped to the 2.3-cM marker interval between C2_At4g17300 and C2_At5g60160 in the introgression. Analysis of a population segregating for Ty-3 and Ty-4 demonstrated that Ty-3 accounted for 59.6% of the variance, while Ty-4 only accounted for 15.7%, suggesting that Ty-4 confers a lesser effect on TYLCV resistance. Recombinant inbred lines (RILs) with Ty-3 and Ty-4 had the highest level of TYLCV resistance. The PCR-based markers tightly linked to the Ty-4 locus as well as the Ty-3 locus have been recently used in our breeding program for efficient selection of high-levels of begomovirus resistance and now allow for efficient breeding by marker-assisted selection.
Tomato-infecting begomoviruses, including monopartite tomato yellow leaf curl virus (TYLCV) and numerous bipartite viruses including tomato mottle virus (ToMoV), are transmitted by the sweetpotato whitefly (Bemisia tabaci), the B biotype of which is also known as the silverleaf whitefly (B. argentifollii). These viruses have caused serious losses to tomato production in many tropical and subtropical regions in the world (Polston and Anderson, 1997). Although the use of insecticides for whitefly control can limit disease, epidemics can still occur, and whitefly resistance to the chemicals has been reported (Omer et al., 1993). Therefore, breeding resistant cultivars would be an effective method for control of this disease. Thus far, TYLCV resistance has not been found in Solanum lycopersicum germplasm, but has been discovered in several tomato wild species, including S. pimpinellifolium, S. peruvianum, S. chilense, S. habrochaites, and S. cheesmaniae (Ji et al., 2007b; Pico et al., 1996; Scott, 2007). Early breeding efforts involved the introgression of resistance genes from the wild accessions into cultivated tomato using traditional breeding approaches (Scott et al., 1996), a process that usually takes many years to breed a resistant cultivar with acceptable horticultural characteristics. Marker-assisted selection (MAS) provides a tool to improve the efficiency of cultivar development. There has been significant progress in the development of markers for important tomato resistance genes (Foolad and Sharma, 2005), including numerous TYLCV resistance genes (Ji et al., 2007b). The first such gene, Ty-1, which originated from S. chilense accession LA1969, was mapped near a molecular marker (TG97) on chromosome 6 of tomato (Zamir et al., 1994). PCR-based markers closely linked to the Ty-1 gene have been developed and can be used for MAS (de Castro et al., 2007; Ji et al., 2007a). A second TYLCV resistance gene, Ty-2 (Hanson et al., 2006), which originated from S. habrochaites (Kalloo and Banerjee, 1990), was mapped to the long arm of chromosome 11, delimited by restriction fragment length polymorphism markers TG393 and TG36 (Hanson et al., 2000). This gene was further delimited to a 4.5-cM marker interval (Ji et al., 2009). We recently identified a third begomovirus resistance gene, Ty-3, in S. chilense accessions LA1932- and LA2779-derived resistant lines, the former carrying a 6-cM introgression and the latter ≈27-cM on the long arm of chromosome 6. Ty-3 was mapped to the same marker interval in the two introgressions, ≈15 cM away from Ty-1 (Ji et al., 2007a). Ty-3 differs from the first two genes mainly in two aspects. First, unlike Ty-1 and Ty-2, which are effective against only monopartite TYLCV, Ty-3 is effective against TYLCV and the bipartite begomovirus ToMoV (Agrama and Scott, 2006; Ji et al., 2007a). In addition, Ty-1 and Ty-2 genes express complete or nearly complete dominance (Hanson et al., 2000; Ji et al., 2009; Zamir et al., 1994), while Ty-3 is more additive (Ji et al., 2007a). Ty-3 is a major locus explaining a high degree of the observed phenotypic variance from the two accessions, but additional resistance loci are required to obtain the highest levels of resistance (Ji et al., 2007a). In a previous study of TYLCV and ToMoV resistance in the backcross progenies derived from various S. chilense accessions, Scott et al. (1996) found multigenic control of resistance derived from S. chilense accessions. Inheritance studies and QTL analysis also demonstrated involvement of at least two genetic loci on chromosome 6 for TYLCV or ToMoV resistance (Agrama and Scott, 2006; Griffiths, 1998; Griffiths and Scott, 2001). Besides Ty-3, we have also searched for additional begomovirus resistance loci in the tomato genome by screening advanced resistant breeding lines derived from various S. chilense accessions with molecular markers on all 12 tomato chromosomes. Herein, we report the presence of a new TYLCV resistance locus, Ty-4, in a S. chilense introgression on the long arm of chromosome 3, and the construction of a molecular linkage map of the Ty-4 locus using a population segregating for Ty-3 and Ty-4 loci.
Materials and Methods
Plant materials.
Four advanced breeding lines derived from each S. chilense accessions LA2779, LA1932, and the intercrosses of lines derived from these two sources (LA2779/LA1932) that displayed a high level of resistance to TYLCV and ToMoV were screened with over 400 PCR-based markers on the 12 tomato chromosomes to identify S. chilense introgressions in tomato genome. These lines were produced from interspecific crosses between tomato and the wild accessions, followed by four to six generations of backcrossing to tomato and six to 10 cycles of self-pollination. Selection for TYLCV and/or ToMoV resistance for over 15 years was done phenotypically by inoculation with one or the other of these pathogens, without any assistance from molecular markers before 2005. In addition to the advanced breeding lines, we also used several early breeding lines (less than five backcrosses) in the screening process, including 960719, 960729, and 960744, which were used as resistant parents in previous studies (Agrama and Scott, 2006; Griffiths, 1998; Griffiths and Scott, 2001). All three were the same for the first three backcrosses (BC3); the latter had a different parent in the fourth BC (BC4), while the former two had the same BC4 parent but diverged in the BC4F2 generation. S. lycopersicum cv. Horizon (Scott et al., 1985) was used as the susceptible control (mean disease severity ≈3.5; see “Inoculation and Disease Evaluation” below), and S. chilense accessions LA2779 and LA1932 [mean disease severity ≈0 (Scott et al., 1996)] were included as the resistant controls for all PCR experiments. An F7 line, 040980, derived from a cross between susceptible S. lycopersicum line 7655B and a begomovirus resistant line 000529, has accessions LA2779 and LA1932 in its pedigree. A population of 201 progeny from selection 040980–3 segregating for TYLCV resistance was employed to map a new TYLCV resistance locus Ty-4.
Plant materials used in a replicated trial to analyze the effects on TYLCV resistance by combining Ty-3 and Ty-4 loci included RILs derived from LA2779/LA1932 containing Ty-3 alone or both Ty-3 and Ty-4, susceptible control ‘Horizon’, and resistant controls including one LA2779-derived breeding line 8602 and two commercial F1 hybrid cultivars Tygress (Seminis, St. Louis, MO) and SecuriTY 28 (Harris Moran, Modesto, CA). Both of these hybrids are heterozygous for the Ty-1 gene. The wild S. chilense accessions were obtained from Tomato Genetics Resource Center at University of California, Davis.
Inoculation and disease evaluation.
The inoculation procedure was the same as described by Griffiths and Scott (2001). In this procedure, seedlings are inoculated with whiteflies from a colony maintained on tomato plants that are infected with TYLCV and thus the whiteflies are viruliferous for this virus. All individual plants were rated for TYLCV disease severity three times according to the method described by Ji et al. (2007a). The third rating was used for statistical analysis to record the most accurate resistance levels of plants and to minimize the possibility of escapes (Griffiths, 1998). The rating scale was from 0 to 4, as documented by Scott et al. (1996), where 0 = no symptoms, 1 = slight symptoms visible only on close inspection, 2 = intermediate symptoms visible on part of the plant, 3 = severe symptoms over the entire plant, and 4 = severe symptoms and stunting. Intermediate scores (0.5, 1.5, etc.) were incorporated to allow for more precise disease severity ratings.
Field experimental design and statistical analysis.
Following inoculation, seedlings were transplanted to field plots at Gulf Coast Research and Education Center, University of Florida, Wimauma. The 201 progeny from selection 040980–3 were transplanted in one continuous block. The seven homozygous RILs and susceptible and resistant control were transplanted to the field in a randomized complete block design with three blocks and 12 plant plots. The soil in the field was Myakka, Haplaquents, and St. Johns sandy soil. The beds were raised 25 cm high, 71 cm wide at the top, and 81 cm wide at the base. Plants were spaced 46 cm apart within plots that were 92 cm apart in rows with 152 cm between rows. The beds had been fumigated with 67% methyl bromide: 33% chloropicrin at 197 kg·ha−1 and the beds were covered with white plastic mulch. The plants were fertilized with a total of 169 kg·ha−1 of nitrogen, 37 kg·ha−1 of phosphorus, and 210 kg·ha−1 of potassium throughout the season through drip irrigation. The plants were staked and tied, and sprayed with pesticides as needed throughout the season according to Olson et al. (2008).
PCR, linkage analysis, and QTL mapping.
All markers used in this study were PCR-based, including sequence-characterized amplified region (SCAR) and cleaved amplified polymorphic sequence (CAPS) markers taken from the public domain or designed from public sequences (Table 1). PCR, linkage analysis, and QTL mapping followed the procedures described by Ji et al. (2007a). The program MAPMAKER/QTL 1.1 (Lincoln et al., 1993) was applied to identify and map the resistance loci and also obtain estimates of the percentage of phenotypic variation explained by each locus as well as components of variance for each resistance locus. Analysis of variance and/or Duncan's multiple range test in SAS (version 9; SAS Institute, Cary, NC) were used to analyze the association of molecular markers with disease resistance in the population segregating for TYLCV resistance, and the TYLCV resistance in the replicated trial of RILs.
Sequence-characterized amplified region (SCAR) and cleaved amplified polymorphic sequence (CAPS) markers on tomato chromosome 3 used to map the Ty-4 locus and near the sp gene on chromosome 6.
Results
S. chilense introgressions in TYLCV-resistant breeding lines.
TYLCV-resistant breeding lines derived from S. chilense accessions LA1932, LA2779, and their intercrosses (LA2779/LA1932) were screened with over 400 molecular markers from the tomato genome to identify introgressions that may carry other TYLCV resistance loci in addition to the Ty-3 locus reported previously (Ji et al., 2007a). Prior information indicated possible S. chilense introgressions on tomato chromosomes 11, 12, 3, and 7 (Y. Ji and J. Scott, unpublished data; Zamir et al. 1994). Therefore, we initially screened the resistant breeding lines with molecular markers on these four chromosomes. The early LA1932-derived line 960719 was shown to carry two additional S. chilense introgressions in addition to the introgression in the Ty-3 region reported previously (Ji et al., 2007a): one is ≈35 cM, spanning markers from TG472 to the sp gene and TG275 on the long arm of chromosome 6, and the other is ≈14 cM, spanning markers from C2_At1g02140 to TG599 on the long arm of chromosome 3 (Fig. 1). Molecular marker analysis also indicated that the early LA1932-derived line 960729 carries only the latter additional introgression besides the Ty-3 introgression. Advanced breeding lines derived from LA2779/LA1932, including line 040980 used for segregation analysis, carry the same introgressions on chromosomes 3 and 6 as did their early ancestor line 090729. Molecular marker analysis in the present study indicated that both introgressed segments in these lines originated from LA1932. Other regions of the tomato genome were also screened with molecular markers, with an average of one marker every 5 cM, but no S. chilense introgression was detected in these regions (data not shown).
One introgressed segment and two separate introgressed segments were identified on chromosomes 3 and 6 of tomato, respectively, in the early breeding lines derived from Solanum chilense accession LA1932, such as lines 960719 (shown) and 960744. The upper smaller introgression (6 cM in length) on chromosome 6 was also present in the advanced resistant lines derived from LA1932, while the lower larger introgression (≈35 cM in length) was not found in the advanced lines for a limited number of markers tested. The introgression (≈14 cM) on chromosome 3 was also present in the advanced resistant breeding lines derived from LA1932 combined with LA2779 (LA2779/LA1932).
Citation: Journal of the American Society for Horticultural Science J. Amer. Soc. Hort. Sci. 134, 2; 10.21273/JASHS.134.2.281
One introgressed segment and two separate introgressed segments were identified on chromosomes 3 and 6 of tomato, respectively, in the early breeding lines derived from Solanum chilense accession LA1932, such as lines 960719 (shown) and 960744. The upper smaller introgression (6 cM in length) on chromosome 6 was also present in the advanced resistant lines derived from LA1932, while the lower larger introgression (≈35 cM in length) was not found in the advanced lines for a limited number of markers tested. The introgression (≈14 cM) on chromosome 3 was also present in the advanced resistant breeding lines derived from LA1932 combined with LA2779 (LA2779/LA1932).
Citation: Journal of the American Society for Horticultural Science J. Amer. Soc. Hort. Sci. 134, 2; 10.21273/JASHS.134.2.281
One introgressed segment and two separate introgressed segments were identified on chromosomes 3 and 6 of tomato, respectively, in the early breeding lines derived from Solanum chilense accession LA1932, such as lines 960719 (shown) and 960744. The upper smaller introgression (6 cM in length) on chromosome 6 was also present in the advanced resistant lines derived from LA1932, while the lower larger introgression (≈35 cM in length) was not found in the advanced lines for a limited number of markers tested. The introgression (≈14 cM) on chromosome 3 was also present in the advanced resistant breeding lines derived from LA1932 combined with LA2779 (LA2779/LA1932).
Citation: Journal of the American Society for Horticultural Science J. Amer. Soc. Hort. Sci. 134, 2; 10.21273/JASHS.134.2.281
Mapping of the TYLCV resistance loci.
Analysis of variance for the population segregating for introgressions on chromosomes 3 and 6 (Ty-3 region) indicated that all the four molecular markers contained within the S. chilense introgression on chromosome 6 were significantly associated with disease severity ratings. For example, the SCAR marker P6–25 designed from bacterial artificial chromosome (BAC) clone 56B23 displayed the highest association with the TYLCV disease severity ratings (Table 2). Similarly, all 12 markers contained within the S. chilense introgression on chromosome 3, such as C2_At4g17300, a CAPS marker derived from a cosII marker, were also significantly associated with disease severity ratings, but to a lesser degree compared with markers from chromosome 6 (Table 2). There was no significant interaction between the genotypes of the two markers P6–25 and C2_At4g17300 (Table 2). However, genotypes with S. chilene alleles at both marker loci displayed the highest level of resistance, while genotypes with S. lycopersicum alleles at both loci showed the severest TYLCV disease (Table 3).
Analysis of variance for the association between markers C2_At4g17300 (linked to Ty-4) on tomato chromosome 3 or P6–25 (linked to Ty-3) on chromosome 6 and the average disease severity of progenies from a plant heterozygous for both Solanum chilense introgressions on chromosome 3 and 6, respectively, which were inoculated with tomato yellow leaf curl virus (TYLCV).
Tomato yellow leaf curl virus disease severity for different combinations of genotypes for markers C2_At4g17300 (linked to Ty-4) on tomato chromosome 3 or P6–25 (linked to Ty-3) on chromosome 6.
Ty-3 was mapped to the marker interval between cLEG-31-P16 (20 cM) and C2_At5g41480 (26 cM) on the long arm of chromosome 6 (Fig. 2). A new TYLCV resistance locus, Ty-4, was mapped in the present study to the marker interval between C2_At4g17300 (81 cM) and C2_At5g60160 (83.3 cM) on the long arm of chromosome 3. Ty-3 and Ty-4 loci had more dominance than additivity, with dominance-to-additive ratios of 0.70 and 1.06, respectively (Table 4). About 60% of the variance in the TYLCV resistance in the population was explained by the Ty-3 locus, while the Ty-4 locus accounted for only ≈16% of the variance, suggesting Ty-3 had a major effect on resistance, while Ty-4 had a lesser effect.
Genetic map of the begomovirus resistance locus Ty-4 on tomato chromosome 3 and the previously mapped begomovirus resistance locus Ty-3 on chromosome 6. Linkage maps were constructed from a mapping population derived from a genotype heterozygous for introgressions on chromosomes 3 and 6 (in the middle). Included on the left is a linkage map for chromosome 3 (designated as Chr. 3) and on the right for chromosome 6 (designated as Chr. 6), which were simplified from the Tomato-EXPEN 2000 map (SGN, 2008). All markers are PCR-based, including sequence-characterized amplified region (SCAR) markers and cleaved amplified polymorphic sequence (CAPS) markers taken from the public domain or designed from public sequences. Shaded regions represent introgressions from Solanum chilense. The markers in nonintrogression regions are not drawn to scale. The numbers on the left of each map represent the published map locations (cM) for the markers (in the Chr. 6 or Chr. 3 map) or linkage distances (cM) between markers in the introgressions obtained from the mapping population.
Citation: Journal of the American Society for Horticultural Science J. Amer. Soc. Hort. Sci. 134, 2; 10.21273/JASHS.134.2.281
Genetic map of the begomovirus resistance locus Ty-4 on tomato chromosome 3 and the previously mapped begomovirus resistance locus Ty-3 on chromosome 6. Linkage maps were constructed from a mapping population derived from a genotype heterozygous for introgressions on chromosomes 3 and 6 (in the middle). Included on the left is a linkage map for chromosome 3 (designated as Chr. 3) and on the right for chromosome 6 (designated as Chr. 6), which were simplified from the Tomato-EXPEN 2000 map (SGN, 2008). All markers are PCR-based, including sequence-characterized amplified region (SCAR) markers and cleaved amplified polymorphic sequence (CAPS) markers taken from the public domain or designed from public sequences. Shaded regions represent introgressions from Solanum chilense. The markers in nonintrogression regions are not drawn to scale. The numbers on the left of each map represent the published map locations (cM) for the markers (in the Chr. 6 or Chr. 3 map) or linkage distances (cM) between markers in the introgressions obtained from the mapping population.
Citation: Journal of the American Society for Horticultural Science J. Amer. Soc. Hort. Sci. 134, 2; 10.21273/JASHS.134.2.281
Genetic map of the begomovirus resistance locus Ty-4 on tomato chromosome 3 and the previously mapped begomovirus resistance locus Ty-3 on chromosome 6. Linkage maps were constructed from a mapping population derived from a genotype heterozygous for introgressions on chromosomes 3 and 6 (in the middle). Included on the left is a linkage map for chromosome 3 (designated as Chr. 3) and on the right for chromosome 6 (designated as Chr. 6), which were simplified from the Tomato-EXPEN 2000 map (SGN, 2008). All markers are PCR-based, including sequence-characterized amplified region (SCAR) markers and cleaved amplified polymorphic sequence (CAPS) markers taken from the public domain or designed from public sequences. Shaded regions represent introgressions from Solanum chilense. The markers in nonintrogression regions are not drawn to scale. The numbers on the left of each map represent the published map locations (cM) for the markers (in the Chr. 6 or Chr. 3 map) or linkage distances (cM) between markers in the introgressions obtained from the mapping population.
Citation: Journal of the American Society for Horticultural Science J. Amer. Soc. Hort. Sci. 134, 2; 10.21273/JASHS.134.2.281
Components of variance for the resistance loci in the mapping population segregating for tomato Ty-3 and Ty-4 loci.
Reduced recombination in the S. chilense introgressed segment.
The recombination length for the whole S. chilense introgressed segment from C2_At1g02140 to TG599 on chromosome 3 in the LA2779/LA1932-derived advanced breeding lines was estimated to be 4.2 cM (Fig. 2), about a 2.5-fold reduction compared with 14 cM in length from the tomato reference map (Fulton et al., 2002; Sol Genomics Network, 2008). Similarly, recombination was greatly reduced in the introgressed segment on chromosome 6, where only one recombinant was found among 201 progeny, giving an estimation of 0.25 cM for the introgression from cLET-31-P16 to C2_At5g41480, which was nearly a 24-fold reduction compared with the same region of 6 cM in length from the tomato reference map.
Enhancing TYLCV resistance by pyramiding Ty-3 and Ty-4.
Analysis of variance for RILs containing Ty-3 alone or in combination with Ty-4 showed significant differences for TYLCV resistance among these lines (Table 5) with no significant difference among the replicates (data not shown). All RILs containing Ty-3 had significantly higher levels of TYLCV resistance than the susceptible controls, as well as the F1 hybrids heterozygous for Ty-1. The two RILs containing Ty-3 and Ty-4 (074766-Y3 and 024525–2) were significantly more resistant than those with Ty-3 alone. Furthermore, plants with homozygous and/or heterozygous Ty-3 and Ty-4 had lower mean disease severity scores than did plants with only Ty-3 in the mapping population (Tables 2 and 3). The level of TYLCV resistance also varied significantly among the four RILs containing Ty-3 without Ty-4 (Table 5), suggesting that additional TYLCV resistance loci besides Ty-3 and Ty-4 may exist in these lines.
Tomato yellow leaf curl virus (TYLCV) disease severity for tomato recombinant inbred lines containing Ty-3 with or without Ty-4.
Discussion
Ty-4, a minor TYLCV resistance locus.
The Ty-3 locus was mapped to the marker interval between cLEG-31-P16 (20 cM) and C2_At5g41480 (26 cM) on the long arm of chromosome 6 from the present investigation, which is consistent with the previous report (Ji et al., 2007a). About 60% of the variance in the TYLCV resistance in the segregating population can be explained by Ty-3 locus, suggesting that Ty-3 is a major locus responsible for TYLCV resistance, a finding also consistent with the previous report (Ji et al., 2007a). The Ty-3 locus was shown to be effective against monopartite TYLCV and bipartite ToMoV (Ji et al., 2007a). In the present study, we showed that Ty-4 accounted for about 16% of the variation and thus would be considered a minor locus compared with Ty-3. An early report also suggested a minor association between an introgression from LA1969 on chromosome 3 and the TYLCV tolerance (Zamir et al., 1994). Ty-4 was effective against TYLCV, but we did not test bipartite begomoviruses. However, inbreds with Ty-3 and Ty-4 have had greater resistance than lines with Ty-3 alone in Guatemala (D.P. Maxwell, unpublished data), suggesting that Ty-4 is effective against multiple (up to seven) bipartite begomoviruses that were reported to be present there (Nakhla et al., 2005).
Combining Ty-3 and Ty-4 loci by MAS.
Plants carrying Ty-3 and Ty-4 loci from the mapping population and RILs showed excellent levels of TYLCV resistance (Tables 2, 3, and 5). These findings support multigenic control of TYLCV resistance in the S. chilense accessions, as observed previously in a study of TYLCV and ToMoV resistance in the backcross progenies derived from various S. chilense accessions in which segregation ratios could not be explained by a single dominant gene (Scott et al., 1996). Inheritance and QTL studies also suggested involvement of at least two loci for TYLCV or ToMoV resistance (Agrama and Scott, 2006; Griffiths and Scott, 2001). The resistance of genotypes homozygous for Ty-3 and Ty-4 was significantly higher than that of genotypes with Ty-3 alone, which in turn were significantly higher than commercial hybrids heterozygous for Ty-1 (Table 5). However, it has been easier to develop commercially acceptable hybrid cultivars using Ty-1 or Ty-2 because they are single dominant genes with high levels of resistance as opposed to using resistance from Ty-3 and/or Ty-4 from S. chilense accessions LA1932 and LA2779, which require more than one gene for the best resistance (Griffiths and Scott, 2001). However, with the discovery of Ty-4 and the development of markers tightly linked to Ty-3 and Ty-4, it is now possible for the first time to use MAS to rapidly incorporate high resistance levels into recurrent parent lines with nearly the same ease as incorporating Ty-1 or Ty-2. One approach would be to put Ty-3 and Ty-4 into both parents of a hybrid. The data in Tables 2 and 3 indicate it might also be possible to have one parent homozygous for both loci while the other could have only Ty-3 or perhaps only Ty-4. Differences in disease severity among the four RILs containing Ty-3 alone suggested additional resistance gene(s) may exist in some LA1932-derived breeding lines. This claim was supported by analysis of TYLCV resistance among the F2 progeny of a LA1932-derived line containing neither Ty-3 nor Ty-4, which segregated for the TYLCV resistance (Y. Ji and J. Scott, unpublished data).
Reduced recombination in the S. chilense introgressions carrying the TYLCV resistance loci.
A nearly 2.5-fold reduction of recombination relative to the tomato reference map (Fulton et al., 2002) was observed in the introgressed segments carrying the Ty-4 locus, which is most likely due to sequence divergence between S. chilense and the cultivated tomato. Reduction of recombination between homeologous chromosomes due to nucleotide divergence was also observed in introgressions from many other tomato wild species (Canady et al., 2006). A genome-wide reduction in recombination frequencies was observed for the introgressed S. lycopersicoides segments in the background of cultivated tomato, often to as low as 0% to 10% of the normal levels (Canady et al., 2006), while recombination within introgressed segments derived from S. pennellii or S. habrochaietesis typically are about 15% to 30% of the normal levels (Alpert and Tanksley, 1996; Monforte and Tanksley, 2000). In the present study, we observed a much greater reduction of recombination (≈24-fold) in the introgressed segments carrying the Ty-3 locus than the 2.5-fold reduction in the introgression carrying the Ty-4 locus. This greater reduction of recombination may be due to the relatively smaller introgressed segment on chromosome 6, which is ≈6 cM in length compared with the 14-cM Ty-4 introgression. This assertion is in agreement with the previous study of a library of S. lycopersicoides introgression lines in which shorter introgressions usually demonstrated greater reductions in homeologous recombination than larger introgressions (Canady et al., 2006).
Reduced recombination within the alien introgressed segments carrying genes of interest may pose potential disadvantages for plant breeders. Plant breeders usually have to make extra efforts (such as more time and larger selecting populations) to break down linkage drag and remove the excessive alien chromatin that causes detrimental effects on the horticultural features of cultivars. Additionally, reduced recombination within introgressed segments may make it difficult to combine tightly linked resistance genes from different sources (in trans configuration) into a single inbred parent (in cis orientation). This is the case for the root-knot nematode resistance Mi gene and the TYLCV resistance gene Ty-1 (Ji et al., 2007b). Numerous tomato breeding programs around the world tried for many years to bring these two genes together in coupling phase, but this was achieved only recently by Seminis Inc. with the assistance of molecular markers (Hoogstraten and Braun, 2005).
Pyramiding resistance genes of various origins to expand the level and range of TYLCV resistance.
The linkage drag situation, however, could be significantly improved using molecular markers tightly linked to genes of interest in the introgressed segments because these markers could facilitate the identification of recombinants with shorter introgressions but still carrying the target gene(s) in succeeding generations. Similarly, with the assistance of molecular markers, it is possible to systematically incorporate multiple resistance genes of different origins into a single elite genotype to enhance the degree of resistance or tolerance to major pathogens such as begomoviruses. Furthermore, pyramiding resistance genes originating from different sources could also expand the resistance against a wider range of begomoviruses (Vidavski, 2007).
Currently, four TYLCV resistance loci (Ty-1 through Ty-4) have been mapped to different regions of the tomato genome, each of which can be tagged with tightly linked PCR-based molecular markers that have been developed in previous and current studies. These resistance loci originate from different wild tomato accessions and possess various modes of TYLCV resistance. Ty-1, which originated from S. chilense accession LA1969 and is nearly completely dominant to TYLCV, is not effective against ToMoV, a bipartite begomovirus (Ji et al., 2007b; Zamir et al., 1994). Ty-2, which originated from S. habrochaites (Kalloo and Banerjee, 1990) and showed completely dominant TYLCV inheritance, is also not effective against ToMoV (Hanson et al., 2000; Ji et al., 2007b). Neither gene is effective against all strains of TYLCV. Unlike Ty-1 and Ty-2, the more recently mapped begomovirus resistant locus Ty-3, which is found in several S. chilense accessions such as LA2779 and LA1932, is more additive in nature and is effective against TYLCV and ToMoV (Agrama and Scott, 2006; Ji et al., 2007a). Ty-4 originated from S. chilense accession LA1932 and has a smaller effect than Ty-3. Robust PCR-based markers developed from current and previous studies such as JB-1, T0302, P6–25, and C2_At4g17300 can be used to effectively tag Ty-1 though Ty-4 loci, respectively, and thus enhance selection efficiency (de Castro et al., 2007; Garcia et al., 2007; Ji et al., 2007c; Ji et al., 2009). In addition, these PCR-based markers can expedite the process of pyramiding these resistance loci into a single elite genotype, thus elevating the level of resistance as well as broadening the range of begomovirus resistance. Lines with different resistance gene combinations need to be tested against a range of begomoviruses to determine their effects, as they can be unpredictable (Vidavski, 2007). Also, the effects of linkage drag may well increase as genes are pyramided and this will have to be addressed.
Literature Cited
Agrama, H.A. & Scott, J.W. 2006 Quantitative trait loci for tomato yellow leaf curl virus and tomato mottle virus resistance in tomato J. Amer. Soc. Hort. Sci. 131 267 272
Alpert, K.B. & Tanksley, S.D. 1996 High-resolution mapping and isolation of a yeast artificial chromosome contig containing fw2.2: A major fruit weight quantitative trait locus in tomato Proc. Natl. Acad. Sci. USA 93 15503 15507
Canady, M.A. , Ji, Y. & Chetelat, R.T. 2006 Homeologous recombination in Solanum lycopersicoides introgression lines of cultivated tomato Genetics 174 1775 1788
de Castro, A.P. , Blanca, J.M. , Díez, M.J. & Viñals, F.N. 2007 Identification of a CAPS marker tightly linked to the tomato yellow leaf curl disease resistance gene Ty-1 in tomato Eur. J. Plant Pathol. 117 347 356
Foolad, M.R. & Sharma, A. 2005 Molecular markers as selection tools in tomato breeding Acta Hort. 695 225 240
Francis, D.M. 2009 PCR-based markers that are polymorphic between Solanum esculentum and S. pimpinellifolium accession LA1589 8 Mar. 2009 <http://164.107.85.47:8003/cgi-bin/molecular_markers.pl>.
Fulton, T.M. , Van der Hoeven, R. , Eannetta, N.T. & Tanksley, S.D. 2002 Identification, analysis, and utilization of conserved ortholog set markers for comparative genomics in higher plants Plant Cell 14 1457 1467
Garcia, B.E. , Graham, E. , Jensen, K.S. , Hanson, P. , Mejía, L. & Maxwell, D.P. 2007 Co-dominant SCAR marker for detection of the begomovirus-resistance Ty-2 locus derived from Solanum habrochaites in tomato germplasm Rpt. Tomato Genet. Coop. 57 21 24
Griffiths, P.D. 1998 Inheritance and linkage of geminivirus resistance genes derived from Lycopersicon chilense (Dunal) in tomato (Lycopersicon esculentum Mill) Ph.D. Diss University of Florida Gainesville
Griffiths, P.D. & Scott, J.W. 2001 Inheritance and linkage of tomato mottle virus resistance genes derived from Lycopersicon chilense accession LA 1932 J. Amer. Soc. Hort. Sci. 126 462 467
Hanson, P.M. , Bernacchi, D. , Green, S. , Tanksley, S.D. , Muniyappa, V. , Padmaja, A.S. , Chen, H. , Kuo, G. , Fang, D. & Chen, J. 2000 Mapping a wild tomato introgression associated with tomato yellow leaf curl virus resistance in a cultivated tomato line J. Amer. Soc. Hort. Sci. 15 15 20
Hanson, P.M. , Green, S.K. & Kuo, G. 2006 Ty-2, a gene on chromosome 11 conditioning geminivirus resistance in tomato Rpt. Tomato Genet. Coop. 56 17 18
Hoogstraten, J.G. & Braun, C.J. 2005 Methods for coupling resistance alleles in tomato 10 Mar. 2009 <http://www.wipo.int/pctdb/en/wo.jsp?wo=2005079342>.
Ji, Y. , Schuster, D.J. & Scott, J.W. 2007a Ty-3, a begomovirus resistance locus near the tomato yellow leaf curl virus resistance locus Ty-1 on chromosome 6 of tomato Mol. Breed. 20 271 284
Ji, Y. , Scott, J.W. & Schuster, D.J. 2009 Toward fine mapping of the tomato yellow leaf curl virus resistance gene Ty-2 on chromosome 11 of tomato HortScience in press
Ji, Y. , Scott, J.W. , Hanson, P. , Graham, E. & Maxwell, D.P. 2007b Sources of resistance, inheritance, and location of genetic loci conferring resistance to members of the tomato-infecting begomoviruses 343 362 Czosnek H. Tomato yellow leaf curl virus disease: Management, molecular biology, breeding for resistance Kluwer Dordrecht, The Netherlands
Ji, Y. , Salus, M.S. , van Betteray, B. , Smeets, J. , Jensen, K.S. , Martin, C.T. , Mejía, L. , Scott, J.W. , Havey, M.J. & Maxwell, D.P. 2007c Co-dominant SCAR markers for detection of the Ty-3 and Ty-3a loci from Solanum chilense at 25 cM of chromosome 6 of tomato Rpt. Tomato Genet. Coop. 57 25 28
Kalloo, G. & Banerjee, M.K. 1990 Transfer of tomato leaf curl virus resistance from Lycopersicon hirsutum f. glabratum to L. esculentum Plant Breed. 105 156 159
Lincoln, S. , Daly, M. & Lander, E.S. 1993 Mapping genes controlling quantitative traits with MAPMAKER/QTL 1.1: A tutorial and reference manual Whitehead Inst. Tech. Rpt. 2nd ed Whitehead Institute Cambridge, MA
Maxwell, D.P. 2009 Introgression at 81 cM on chromosome 3 associated with Gc171 8 Mar. 2009 <http://www.plantpath.wisc.edu/GeminivirusResistantTomatoes/Markers/P3-81.pdf>.
Monforte, A.J. & Tanksley, S.D. 2000 Fine mapping of a quantitative trait locus (QTL) from Lycopersicon hirsutum chromosome 1 affecting fruit characteristics and agronomic traits: Breaking linkage among QTLs affecting different traits and dissection of heterosis for yield Theor. Appl. Genet. 100 471 479
Nakhla, M.K. , Sorenson, A. , Mejía, L. , Ramírez, P. , Karkashian, J.P. & Maxwell, D.P. 2005 Molecular characterization of tomato-infecting begomoviruses in Central America and development of DNA-based detection methods Acta Hort. 695 277 288
Olson, S.M. , Maynard, D.N. , Hochmuth, G.J. , Vavrina, C.S. , Stall, W.M. , Kucharek, T.A. , Webb, S.E. , Taylor, T.G. , Smith, S.A. & Simonne, E. 2008 Tomato production in Florida 409 429 Olson S.M. & Simonne E. Vegetable production handbook for Florida 2007–2008 Vance Publishing Lincolnshire, IL
Omer, A.D. , Johnson, M.W. , Tabashnik, B.E. , Costa, H.S. & Ullman, D.E. 1993 Sweet-potato whitefly resistance to insecticides in Hawaii: Intra-island variation is related to insecticide use Entomol. Exp. Appl. 67 173 182
Pico, B. , Diez, M.J. & Nuez, F. 1996 Viral diseases causing the greatest economic losses to the tomato crop. II. The tomato yellow leaf curl virus: A review Scientia Hort. 67 151 196
Polston, J.E. & Anderson, P.K. 1997 The emergence of whitefly-transmitted geminiviruses in tomato in the western hemisphere Plant Dis. 81 1358 1369
Scott, J.W. 2007 Breeding for resistance to viral pathogens 447 474 Razdan M.K. & Mattoo A.K. Genetic improvement of solanaceous crops. Vol. 2: Tomato Science Publisher Enfield, NH
Scott, J.W. , Bartz, J.A. , Bryan, H.H. , Everett, P.H. , Gull, D.D. , Howe, T.K. , Stoffella, P.J. & Volin, R.B. 1985 ‘Horizon’: A fresh market tomato with concentrated fruit set Florida Agr. Expt. Sta. Circ. S-323 8
Scott, J.W. , Stevens, M.R. , Barten, J.H.M. , Thome, C.R. , Polston, J.E. , Schuster, D.J. & Serra, C.A. 1996 Introgression of resistance to whitefly-transmitted geminiviruses from Lycopersicon chilense to tomato 357 367 Gerling D. & Mayer R.T. Bemisia 1995: Taxonomy, biology, damage control, and management Intercept Andover, UK
Sol Genomics Network 2008 Tomato-EXPEN 2000: S. lycopersicum LA925 × S. pennellii LA716 type F2.2000 18 Dec. 2008 <http://www.sgn.cornell.edu/cview/map.pl?map_id=9&show_offsets=1&show_ruler=1>.
Vidavski, F.S. 2007 Exploitation of resistance genes found in wild tomato species to produce resistant cultivars; pile up of resistant genes 363 372 Czosnek H. Tomato yellow leaf curl virus disease: Management, molecular biology, breeding for resistance Kluwer Dordrecht, The Netherlands
Zamir, D. , Eksteinmichelson, I. , Zakay, Y. , Navot, N. , Zeidan, M. , Sarfatti, M. , Eshed, Y. , Harel, E. , Pleban, T. , Vanoss, H. , Kedar, N. , Rabinowitch, H.D. & Czosnek, H. 1994 Mapping and introgression of a tomato yellow leaf curl virus tolerance gene, Ty-1 Theor. Appl. Genet. 88 141 146
