Non-transgenic Tomato Research Articles

Knockdown of orthotospovirus-derived silencing suppressor gene by plant-mediated RNAi approach induces viral resistance in tomato

RNA interference (RNAi) shows significant effectiveness in conferring resistance to viral infections in various plants. The current study was conducted to develop transgenic tomato (Solanum lycopersicum cv. Rio Grande) lines expressing a 320 bp conserved inverted region of the integral 35S promoter region. The molecular cloning was done for the construction of expression vectors in a hairpin structure by the Tomato spotted wilt orthotospovirus (TSWV) species Orthotospovirus tomatomaculae, silencing suppressor (NSs) gene. It contains both sense and antisense orientation in a binary vector pFGC5941. Short hairpin RNA (shRNA) structures were employed to reduce the possibility of inducing endogenous non-specific antiviral responses, which usually target longer double-stranded RNAs (dsRNAs). To confirm the transgenic plants, molecular analysis was performed using TSWV gene-specific primers (TSWV-NSs). Based on the EHA105 Agrobacterium strain and the pFGC5941 vector carrying NSs and nptII genes in the T-DNA region, the results validate the insertion of kanamycin resistance into the regenerated transgenic plants. The virus source harboring TSWV was used to inoculate putative transgenic plants for evaluating the confirmation of successful transformation strategy for inducing virus resistance. Quantitative real-time polymerase chain reaction (qRT-PCR) assays were performed by using qRT-NSs primer pairs to determine the relative gene expression of all constructs (TSWV-NSs both sense and antisense orientation) after inoculations. In comparison to the control groups, which contained wild type control (non-transgenic tomato plant), empty pFGC5941 vector and the pFGC5941+sense construct, the expression of the NSs gene was noticeably reduced in the full clone (pFGC5941+sense + antisense) construct. For the validation of RNAi effectiveness, Nicotiana tabacum plants were also inoculated with viral constructs. Specifically, they displayed severe symptoms of TSWV in plants containing the pFGC5941+sense, pFGC5941 empty vector, and in wild-type tomato plants that are correlated with bioassay results. The lack of TSWV symptoms validated the hypothesis after careful observation of the expression of NSs genes in terms of pathogenicity. This research demonstrates the RNA interference effectiveness by targeting NSs gene as a reliable technique for achieving long lasting antiviral protection in tomatoes. The findings have practical relevance and the potential to be a tool for limiting tomato crop losses caused by TSWV worldwide.

The comparative analysis of agronomic, compositional, and physiological traits of miraculin transgenic tomato in the confined field trial

<abstract> <p>The miraculin transgenic tomato is a genetically modified (GM) crop that can be used as an alternative for low calories food and a natural non-sugar sweetener. Before the release and distribution, transgenic crop needs to go through an environmental risk assessment (ERA) as a backbone to achieve biosafety. Comparative analysis is a general principle of ERA to identify differences between transgenic crop and its non-transgenic counterpart which may indicate substantial equivalence and unintended effects. This experiment was aimed to compare the agronomic, compositional, and physiological characteristics of miraculin transgenic tomato cv. Moneymaker with non-transgenic tomato. The data obtained were plant height, stem diameter, relative growth rate, chlorophyll content, stomatal conductance, days to 50% flowering, days to fruit maturity, a number of flowers per cluster, a number of fruits per cluster, a number of fruits per plant, fruit weight, fruit diameter, harvest index, total dissolved solids, fructose, glucose, and sucrose contents, and total carotenoids, lycopene, and β-carotene contents. This study found that there were no significant differences between miraculin transgenic and non-transgenic tomato in all variables observed. It suggests that miraculin transgenic tomato is equivalent to its counterpart and unintended effects are not detected as.</p> </abstract>

Evaluation of agronomic and fruit quality traits of miraculin transgenic tomato

Abstract. Carsono N, Desiana N, Nurrizqi FM, Elfakhriano IF, Anas, Sari S, Kusumiyati, Ohsawa R, Shimono A, Ezura H. 2022. Evaluation of agronomic and fruit quality traits of miraculin transgenic tomato. Biodiversitas 23: 2004-2009. Tomato cv. Moneymaker has been inserted with miraculin gene which can modify sour taste becomes sweet in the human tongue and, thus, it has potential as a sweetener. Evaluation of agronomic and fruit quality traits for this special transgenic tomato is a major concern that should be performed as substantial equivalence. The study's objective was to compare agronomic and fruit quality traits of transgenic miraculin tomato with those of its wild-type. The experiment employed a randomized block design, one genotype factor with two levels (transgenic vs. non-transgenic tomato cv. Moneymaker), and each was four replicates. Data were subjected to the Student t-test. The study revealed no significant differences between the transgenic tomato plants cv. Moneymaker and the wild type for important agronomic traits such as plant height, stem diameter, leaf area index, fruit weight, fruit weight per cluster, fruit weight per plant, fruit diameter, number of fruits per cluster, number of fruits per plant, and harvest index of miraculin. Some fruit quality traits such as fruit shelf-life, fruit hardness, total soluble solids, and content of glucose, sucrose, and fructose were equivalent to those of its counterpart, suggesting that miraculin transgene did not affect the expression of the observed traits. The miraculin transgenic tomato traits were more likely identical to those of wild-type except for the existence of the miraculin transgene. This will open the possibility for this special transgenic tomato to undergo further environmental safety assessment in the field test facility as a part of the biosafety assessment of genetically modified organism (GMO).

Invasiveness, allelopathic potential and unintended effects of miraculin transgenic tomato to soil microbes

<abstract> <p>Tomato cv. Moneymaker was modified by the insertion of a miraculin gene, which can modify a sour taste into a sweet taste. Environmental safety assessment for this special transgenic crop is an important step in assessing how safe this tomato is before it is released into the environment. Evaluation of invasiveness, allelopathy and unintended effects is highly essential for environmental safety assessment. The evaluation of invasiveness was carried out by growing a mixture of transgenic and non-transgenic tomatoes with ratios of 0:100 and 100:0 (sole-cultivation) and 25:75, 50:50 and 75:25 (mix-cultivation). Wet and dry biomasses of three-week-old tomato plants were measured. Soil microbes were evaluated by determining microbial populations (culturable) and estimating soil respiration. Microbial populations were determined through total plate count, while soil respiration was estimated using the titration method to calculate the levels of carbon dioxide released during the incubation. It was found that the aggressiveness of the miraculin transgenic tomato was equal to that of its counterpart. There were also no significant differences in microbial populations and soil respiration of miraculin transgenic tomato compared with those of wild type. In addition, miraculin transgenic tomato did not produce allelopathy that interfered with surrounding crops. It is concluded that transgenic tomato is equal to its counterpart in invasiveness, with no effect to soil microbes and no potential allelopathy found.</p> </abstract>

MtNOOT gene enhanced high productivity and economical characteristics of tomato (Lycopersicon esculentum)

Tomato (Lycopersicon esculentum) is the main vegetal crop that has tremendous popularity around the world. Medicago truncatula NOOT gene (Mt-NOOT) encodes a BTB/POZ-ankyrin repeat protein of the NONEXPRESSOR OF PR GENES1 (NPR1 family). It was introduced into Lycopersicon esculentum (Tomato) genome. The tomato plants that ectopically expressed Mt-NOOT obtained several favorable traits and fruit quality. Heteroblasty between the transgenic and the non-transgenic tomato leaves and flower architecture were used to distinguish transgenic and wild lines. Transgenic tomato plants accumulated a significant amount of phenolic compounds and plant pigmentations compared to the wild type. On the other hand, transgenic plants acquired a considerable amount of antioxidant such as CuZnSO superoxide dismutase (SOD), tomato Catalase (CAT), and tomato Cell wall-associated peroxidase (TPX1) than the wild type. Antioxidant high content together with the high content of phenolic compounds enabled the transgenic tomato fruits to gain not only edible benefits, but also a significant higher shelf-time, extended to six months more than the wild type stored at 25°C in dark and dry condition. Surprisingly, transgenic tomato fruits did not show any rotten process during long time storage as they did not acquire any contagious microorganism. Total fruit productivity in transgenic tomato was greater than the control with an estimated ratio of 84%.

Development of zeaxanthin‐rich tomato fruit through genetic manipulations of carotenoid biosynthesis

SummaryThe oxygenated carotenoid zeaxanthin provides numerous benefits to human health due to its antioxidant properties. Especially it is linked to protecting, together with the xanthophyll lutein, the retina in the human eye by filtering harmful blue light thus delaying the progression of age‐related macular degeneration (AMD), the most prevalent cause of blindness in developed countries. Despite its high nutritional value, zeaxanthin is less available than other substantial carotenoids in our diet. To solve this shortage, we chose to develop a new food source that would contain a high concentration of natural zeaxanthin. Tomato (Solanum lycopersicum L.) was selected as the target plant since it is the second largest vegetable crop grown worldwide and its fruit characteristically synthesizes and accumulates a high concentration of carotenoids. We employed two genetic approaches in order to enhance zeaxanthin biosynthesis in tomato fruit: a transgenic metabolic engineering and classical genetic breeding. A nontransgenic tomato line, named ‘Xantomato’, was generated whose fruit accumulated zeaxanthin at a concentration of 39 μg/g fresh weight (or 577 μg/g dry weight), which comprised ca. 50% of total fruit carotenoids compared to zero in the wild type. This is the highest concentration of zeaxanthin reached in a primary crop. Xantomato can potentially increase zeaxanthin availability in the human diet and serve as raw material for industrial applications.

Remusatia vivipara lectin and Sclerotium rolfsii lectin interfere with the development and gall formation activity of Meloidogyne incognita in transgenic tomato.

Root knot nematodes are serious threats to growth and yield of solaneous cropsincludingtomato. In this study, a binary vector carrying Remusatia vivipara (rvl1) and Sclerotium rolfsii (srl1) lectin genes were introduced independently into Lycopersicon esculentum cv. Pusa Ruby via Agrobacterium tumefaciens for resistance against root knot nematode, Meloidogyne incognita. In total, one hundred and one rvl1 and srl1-transformed plants exhibiting kanamycin resistance were confirmed to carry transgenes as detected by polymerase chain reaction (PCR) with 4.59% transformation efficiency. Genetic analysis of T1 progeny confirmed Mendelian segregation of the introduced genes. Three events each of rvl1 and srl1 transgenic tomato were randomly selected for further confirmation by Southern and TAIL-PCR analyses. All three events of srl1 transgenics showed single copy transgene, whereas two rvl1 transgenic events showed single copy of transgene, while remaining event showed two copies of transgenes. Site of integration obtained for rvl1 and srl1 transgenic events by TAIL-PCR revealed that all the three events of rvl1 and srl1 transgenics differed for their site of integration and insertion sites did not contain any predicted gene. Moreover, expression of the rvl1 and srl1 transgenes was detected by haemagglutination assay in all three events of rvl1 and srl1, but not in non-transgenic tomato plant. Homozygous progenies of these events were grown and inoculated with M. incognita. Development and reproduction of M. incognita was severely affected in transgenic tomato plants expressing RVL1 and SRL1 exhibiting the high levels of resistance compared to non-transgenic plants. Therefore, these transgenic lines demonstrate a promising potential for variety development of tomato lines with enhanced resistance against M. incognita.

Rapid generation of a transgene-free powdery mildew resistant tomato by genome deletion

Genome editing has emerged as a technology with a potential to revolutionize plant breeding. In this study, we report on generating, in less than ten months, Tomelo, a non-transgenic tomato variety resistant to the powdery mildew fungal pathogen using the CRISPR/Cas9 technology. We used whole-genome sequencing to show that Tomelo does not carry any foreign DNA sequences but only carries a deletion that is indistinguishable from naturally occurring mutations. We also present evidence for CRISPR/Cas9 being a highly precise tool, as we did not detect off-target mutations in Tomelo. Using our pipeline, mutations can be readily introduced into elite or locally adapted tomato varieties in less than a year with relatively minimal effort and investment.

A novel strategy to enhance resistance to Cucumber mosaic virus in tomato by grafting to transgenic rootstocks

Cucumber mosaic virus (CMV) can infect a wide range of host species. For the lacking of CMV resistant varieties of tomato, RNA interference (RNAi) can be used as a fast and effective method for the generation of transgenic resistant varieties. In this current study, five intron-spliced hairpin RNA (ihpRNA) plant expression vectors aimed at five genes of CMV have been constructed. Transgenic tomatoes were obtained by Agrobacterium tumefaciens-mediated transformation with expression vectors. Highly resistant generations of transgenic plants were employed as rootstocks and grafted onto non-transgenic tomatoes that resulted in the successful transfer of resistance to the scions. Using a novel method of plant cuttings for rootstock propagation, we obtained large quantities of disease-resistant material. Further, this method produces scions that can remain undetectable for transgenic resistance marker genes that may provide novel approaches to evade collective concerns about genetically-modified organism (GMO) biosafety.

Transmission of RNA silencing signal through grafting confers virus resistance from transgenically silenced tobacco rootstocks to non-transgenic tomato and tobacco scions

We examined the transmission of RNA silencing signal in non-transgenic tomato and tobacco scions grafted onto the tobacco Sd1 rootstocks, which is silenced in both NtTOM1 and NtTOM3 required for tobamovirus multiplication. When the non-transgenic tomato scions were grafted onto the Sd1 rootstocks, RT-PCR analysis of the scions showed the reduced level of mRNA compared with that before grafting in both LeTH3 and LeTH1, tomato homologs of NtTOM1 and NtTOM3, respectively. siRNAs from both genes were detected in the scions after grafting but not before grafting. Further tomato scions were inoculated with Tomato mosaic virus (ToMV) and used for virus infection. They showed very low level of virus accumulation. Necrotic responding tobacco to tobamovirus was grafted onto the rootstock of Sdl. RT-PCR analysis showed low level expression of both NtTOM1 and NtTOM3 in the scions but siRNA was detected after grafting. When the leaves of scions were inoculated with ToMV or Tobacco mosaic virus, they produced very few local necrotic lesions (LNLs) while the control scions did many LNLs. These results suggest that RNA silencing was transmitted to non-transgenic tomato and tobacco scions after grafting onto the Sd1 rootstocks and that virus resistance was induced in the scions.

A novel strategy to produce sweeter tomato fruits with high sugar contents by fruit‐specific expression of a single bZIP transcription factor gene

Enhancement of sugar content and sweetness is desirable in some vegetables and in almost all fruits; however, biotechnological methods to increase sugar content are limited. Here, a completely novel methodological approach is presented that produces sweeter tomato fruits but does not have any negative effects on plant growth. Sucrose-induced repression of translation (SIRT), which is mediated by upstream open reading frames (uORFs), was initially reported in Arabidopsis AtbZIP11, a class S basic region leucine zipper (bZIP) transcription factor gene. Here, two AtbZIP11 orthologous genes, SlbZIP1 and SlbZIP2, were identified in tomato (Solanum lycopersicum). SlbZIP1 and SlbZIP2 contained four and three uORFs, respectively, in the cDNA 5'-leader regions. The second uORFs from the 5' cDNA end were conserved and involved in SIRT. Tomato plants were transformed with binary vectors in which only the main open reading frames (ORFs) of SlbZIP1 and SlbZIP2, without the SIRT-responsive uORFs, were placed under the control of the fruit-specific E8 promoter. Growth and morphology of the resulting transgenic tomato plants were comparable to those of wild-type plants. Transgenic fruits were approximately 1.5-fold higher in sugar content (sucrose/glucose/fructose) than nontransgenic tomato fruits. In addition, the levels of several amino acids, such as asparagine and glutamine, were higher in transgenic fruits than in wild-type fruits. This was expected because SlbZIP transactivates the asparagine synthase and proline dehydrogenase genes. This 'sweetening' technology is broadly applicable to other plants that utilize sucrose as a major translocation sugar.

Conferring high-temperature tolerance to nontransgenic tomato scions using graft transmission of RNA silencing of the fatty acid desaturase gene.

We investigated graft transmission of high-temperature tolerance in tomato scions to nontransgenic scions from transgenic rootstocks, where the fatty acid desaturase gene (LeFAD7) was RNA-silenced. Tomato was transformed with a plasmid carrying an inverted repeat of LeFAD7 by Agrobacterium. Several transgenic lines showed the lower amounts of LeFAD7 RNA and unsaturated fatty acids, while nontransgenic control did not, and siRNA was detected in the transgenic lines, but not in control. These lines grew under conditions of high temperature, while nontransgenic control did not. Further, the nontransgenic plants were grafted onto the silenced transgenic plants. The scions showed less of the target gene RNA, and siRNA was detected. Under high-temperature conditions, these grafted plants grew, while control grafted plants did not. Thus, it was shown that high-temperature tolerance was conferred in the nontransgenic scions after grafting onto the silenced rootstocks.

RNA interference-based resistance in transgenic tomato plants against Tomato yellow leaf curl virus-Oman (TYLCV-OM) and its associated betasatellite.

BackgroundTomato yellow leaf curl virus (TYLCV), a monopartite begomovirus (family Geminiviridae) is responsible for heavy yield losses for tomato production around the globe. In Oman at least five distinct begomoviruses cause disease in tomato, including TYLCV. Unusually, TYLCV infections in Oman are sometimes associated with a betasatellite (Tomato leaf curl betasatellite [ToLCB]; a symptom modulating satellite). RNA interference (RNAi) can be used to develop resistance against begomoviruses at either the transcriptional or post-transcriptional levels.ResultsA hairpin RNAi (hpRNAi) construct to express double-stranded RNA homologous to sequences of the intergenic region, coat protein gene, V2 gene and replication-associated gene of Tomato yellow leaf curl virus-Oman (TYLCV-OM) was produced. Initially, transient expression of the hpRNAi construct at the site of virus inoculation was shown to reduce the number of plants developing symptoms when inoculated with either TYLCV-OM or TYLCV-OM with ToLCB-OM to Nicotiana benthamiana or tomato. Solanum lycopersicum L. cv. Pusa Ruby was transformed with the hpRNAi construct and nine confirmed transgenic lines were obtained and challenged with TYLCV-OM and ToLCB-OM by Agrobacterium-mediated inoculation. For all but one line, for which all plants remained symptomless, inoculation with TYLCV-OM led to a proportion (≤25%) of tomato plants developing symptoms of infection. For inoculation with TYLCV-OM and ToLCB-OM all lines showed a proportion of plants (≤45%) symptomatic. However, for all infected transgenic plants the symptoms were milder and virus titre in plants was lower than in infected non-transgenic tomato plants.ConclusionsThese results show that RNAi can be used to develop resistance against geminiviruses in tomato. The resistance in this case is not immunity but does reduce the severity of infections and virus titer. Also, the betasatellite may compromise resistance, increasing the proportion of plants which ultimately show symptoms.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-015-0263-y) contains supplementary material, which is available to authorized users.

Proteomic Analysis of Tomato (Lycopersicum esculentum var. cerasifarm) Expressing the HBsAg Gene by 2-dimensional Difference Gel Electrophoresis

In a previous study, an HBsAg gene-bearing transgenic tomato line was made available and it exhibited notable physiological alterations compared with the non-transgenic tomato (control). In particular, leaves of the transgenic plants were fleshy and dark. We hypothesized that a change in leaf proteins of the transgenic plants account for the observed phenotypes. In this study, total protein content in leaves of the transgenic plants was analyzed by 2-dimensional difference gel electrophoresis. A total number of 700 protein spots were detected on silver-stained gels, of which 368 protein spots were matched between the control and sample gels. Among these matched proteins, the expression levels of 122 proteins in the transgenic plants were upregulated while those of the rest were downregulated. In addition, 25 abundant proteins (value ratio > 2.0) on silver-stained gels were analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Sixteen differentially expressed proteins were identified, of which 13 were predicted to be involved in cell division, energy metabolism, protein synthesis and processing. The possible roles of these proteins in the transgenic tomato strain have been discussed. Taken together, our data indicate that significant alterations in protein expression occur in transgenic tomatoes bearing the HBsAg gene. Our findings will help broaden our knowledge of the mechanism by which exogenously expressed genes lead to phenotypic alterations in transgenic plants.

Rapid determination of ethylene content in tomatoes using visible and short-wave near-infrared spectroscopy and wavelength selection

The plant hormone ethylene controls many aspects of development. In tomato, ethylene is an essential component for fruit ripening. In this paper, the study was concentrated on the visible/short-wave near-infrared (Vis/SW NIR) spectroscopy technique for the quantitative analysis of ethylene content in three varieties of tomatoes: non-transgenic tomatoes, and transgenic tomatoes with antisense LeETR1 and LeETR2. The results indicate that the determination of ethylene content in tomatoes could be successfully performed through Vis/SW NIR spectroscopy combined with chemometrics methods including partial least square regression (PLSR) and stepwise multiple linear regression (SMLR). The performances of models using four spectral pretreatment methods – standard normal variate (SNV), multiplicative scatter correction (MSC), first and second derivation – were compared. Wavelengths for ethylene analysis in tomatoes were proposed using SMLR models. The models of ethylene in tomato using the assigned wavelengths show reliable results. The prediction precision of PLSR and SMLR using selected wavelengths was compared. The results show that the modeling of PLSR using the visible region needed less time than that using full region, although the prediction precisions were a little lower. The overall results indicate that Vis/SW NIR spectroscopy combined with wavelength selection could be applied as a nondestructive and rapid tool for the determination of ethylene content in tomatoes.

Resistance Enhancement of Transgenic Tomato to Bacterial Pathogens by the Heterologous Expression of Sweet Pepper Ferredoxin-I Protein

ABSTRACT Expression of a foreign gene to enhance plant disease resistance to bacterial pathogens is a favorable strategy. It has been demonstrated that expressing sweet pepper ferredoxin-I protein (PFLP) in transgenic plants can enhance disease resistance to bacterial pathogens that infect leaf tissue. In this study, PFLP was applied to protect tomato (Lycopersicon esculentum cv. cherry Cln1558a) from the root-infecting pathogen, Ralstonia solanacearum. Independent R. solanacearum resistant T(1) lines were selected and bred to produce homozygous T(2) generations. Selected T(2) transgenic lines 24-18-7 and 26-2-1a, which showed high expression levels of PFLP in root tissue, were resistant to disease caused by R. solanacearum. In contrast, the transgenic line 23-17-1b and nontransgenic tomato, which showed low expression levels of PFLP in root tissue, were not resistant to R. solanacearum infection. The expansion of R. solanacearum populations in stem tissue of transgenic tomato line 24-18-7 was limited compared with the nontransgenic tomato Cln1558a. Using a detached leaf assay, transgenic line 24-18-7 was also resistant to maceration caused by E. carotovora subsp. carotovora; however, resistance to E. carotovora subsp. carotovora was less apparent in transgenic lines 26-2-1a and 23-17-1b. These results demonstrate that PFLP is able to enhance disease resistance at different levels to bacterial pathogens in individual tissue of transgenic tomato.

Quantification of Chlorophyll Content and Classification of Nontransgenic and Transgenic Tomato Leaves Using Visible/Near-Infrared Diffuse Reflectance Spectroscopy

Visible/near-infrared (vis/NIR) spectroscopy combined with multivariate analysis was used to quantify chlorophyll content in tomato leaves and classify tomato leaves with different genes. In this study, transgenic tomato leaves with antisense LeETR1 (n = 106) and their parent nontransgenic ones (n = 102) were measured in vis/NIR diffuse reflectance mode. Quantification of chlorophyll content was achieved by partial least-squares regression with a cross-validation prediction error equal to 2.87. Partial least-squares discriminant analysis was performed to classify leaves. The results show that differences between transgenic and nontransgenic tomato leaves do exist, and excellent classification can be obtained after optimizing spectral pretreatment. The classification accuracy can reach to 100% using the derivative of spectral data in the full and partial wavenumber range. These results demonstrate that vis/NIR spectroscopy together with chemometrics techniques could be used to quantify chlorophyll content and differentiate tomato leaves with different genes, which offers the benefit of avoiding time-consuming, costly, and laborious chemical and sensory analysis.

Combination and comparison of chemometrics methods for identification of transgenic tomatoes using visible and near-infrared diffuse transmittance technique

The potential of visible/near-infrared (Vis/NIR) diffuse transmittance spectroscopy, in combination with a variety of chemometrics techniques, was investigated to examine the feasibility to non-destructively distinguish transgenic tomatoes from non-transgenic tomatoes. One hundred tomatoes inserted with antisense ethylene receptor gene LeETR1 and the same number of their parents were scanned in the Vis/NIR regions. Principal component analysis (PCA), soft independent modelling of class analogy (SIMCA) and discriminant partial least squares (DPLS) regression based on PCA scores were applied to classify tomatoes with different genes into two groups. The results show that differences between transgenic and non-transgenic tomatoes do exist and excellent classification can be obtained after optimizing spectral pre-treatment. The correct classifications of the calibration as well as the validation data set for transgenic and non-transgenic tomatoes were 100% using DPLS after second derivative spectral pre-treatment. The results in the present study show that Vis/NIR spectroscopy together with chemometrics techniques could be a rapid tool to be used for differentiating transgenic tomatoes from conventional tomatoes.

Making a friend from a foe: expressing a GroEL gene from the whitefly Bemisia tabaci in the phloem of tomato plants confers resistance to tomato yellow leaf curl virus

Some (perhaps all) plant viruses transmitted in a circulative manner by their insect vectors avoid destruction in the haemolymph by interacting with GroEL homologues, ensuring transmission. We have previously shown that the phloem-limited begomovirus tomato yellow leaf curl virus (TYLCV) interacts in vivo and in vitro with GroEL produced by the whitefly vector Bemisia tabaci. In this study, we have exploited this phenomenon to generate transgenic tomato plants expressing the whitefly GroEL in their phloem. We postulated that following inoculation, TYLCV particles will be trapped by GroEL in the plant phloem, thereby inhibiting virus replication and movement, thereby rendering the plants resistant. A whitefly GroEL gene was cloned in an Agrobacterium vector under the control of an Arabidopsis phloem-specific promoter, which was used to transform two tomato genotypes. During three consecutive generations, plants expressing GroEL exhibited mild or no disease symptoms upon whitefly-mediated inoculation of TYLCV. In vitro assays indicated that the sap of resistant plants contained GroEL-TYLCV complexes. Infected resistant plants served as virus source for whitefly-mediated transmission as effectively as infected non-transgenic tomato. Non-transgenic susceptible tomato plants grafted on resistant GroEL-transgenic scions remained susceptible, although GroEL translocated into the grafted plant and GroEL-TYLCV complexes were detected in the grafted tissues.

Discrimination of transgenic tomatoes based on visible/near-infrared spectra

VIS–NIR spectroscopy combined with multivariate analysis after the appropriate spectral data pre-treatment has been proved to be a very powerful tool for judgment of the relative pattern of the objects that have very similar properties. In this study, seventy transgenic tomatoes with antisense LeETR2 and 94 of their parents, non-transgenic ones were measured in VIS–NIR diffuse reflectance mode. Principal component analysis (PCA), discriminant analysis (DA) and partial least-squares discriminant analysis (PLSDA) were applied to classify tomatoes with different genes into two groups. Calibrations were developed using PLS regression with the leave-one-out cross-validation technique. The results show that differences between transgenic and non-transgenic tomatoes do exist and excellent classification can be obtained after optimizing spectral pre-treatment. The correct classifications for transgenic and non-transgenic tomatoes were both 100% using PLSDA after derivative spectral pre-treatment. The raw spectra with PLSDA model after the second derivative pre-treatment had the best satisfactory calibration and prediction abilities, with r c = 0.97964, root mean square error of calibration (RMSEC) = 0.099, r cv = 0.97963, root mean square error of cross-validation (RMSECV) = 0.0993 and a factor. The results in the present study show VIS–NIR spectroscopy together with chemometrics techniques could be used to differentiate transgenic tomato, which offers the benefit of avoiding time-consuming, costly and laborious chemical and sensory analysis.