番茄S-腺苷蛋氨酸脱羧酶基因SlSAMDC1的克隆与序列分析

 以蔓陀罗S-腺苷蛋氨酸脱羧酶全长cDNA序列为信息探针, 筛选NCBI番茄EST数据库, 依据同源EST信息,经人工拼接、RT-PCR及RACE技术验证,获得了1个新的SAMDC基因家族成员,命名为SlSAMDC1(GenBank登录号：EF550528),并利用染色体步移技术克隆了879 bp的上游调控区域。SlSAMDC1 cDNA序列全长1 847 bp, 5′-UTR和3′-UTR分别长523 bp、241 bp;存在3个ORF(微型uORF、小型uORF和主ORF),主ORF编码360个氨基酸的SAMDC酶原;SlSAMDC1基因组序列全长3 648 bp,有3个内含子,均位于5′-UTR,所有内含子的剪切位点均符合真核生物“GT-AG”规则。SlSAMDC1基因与其它植物来源SAMDC基因同源性较高,与人、大肠杆菌以及酵母的同源性较低。表达谱分析发现,SlSAMDC1基因在番茄根、茎、叶、花蕾、果实等器官中均表达,果实中的表达量相对较高。生物信息学分析表明,SlSMDC1基因的上游调控序列存在多个顺式作用元件,如W-box、TATA-box、CAAT-box等。     

Improvements in somatic embryogenesis protocol in Feijoa (Acca sellowiana (Berg) Burret): Induction,conversion and synthetic seeds

Pineapple guava (Acca sellowiana) syn. Feijoa sellowiana, a Brazilian indigenous Myrtaceae is under domestication in South Brazil. Previous works showed that this species is responsive to somatic embryogenesis and recalcitrant to conventional methods of clonal propagation. In the present work it was evaluated the role of components of culture medium in the induction and development of somatic embryos. The technology of synthetic seeds was also evaluated. Zygotic embryos were inoculated in LPm medium supplemented with 8 mM glutamic acid and 8 mM l-glutamine, 2,4-dichlophenoxiacetic acid (20 μM) and myo-inositol. For conversion of somatic embryos and synthetic seeds it was tested the effect of 6-benzylaminopurine and gibberellic acid combined or not with activated charcoal. The highest values for embryogenetic induction (100%) and number of somatic embryos/explant (113) were observed in the LPm medium supplemented with Glu (8 mM), and 2,4-D. The culture medium supplemented with BA (0.5 μM) and GA₃ (1 μM) and activated charcoal (1.5 g L⁻¹) enhanced the conversion of somatic embryos to plantlets. Pre-germinated somatic embryos encapsulated in sodium alginate with BA (0.5 μM) and GA₃ (1 μM) developed radicles. The use of synthetic seed was a requisite for the survival of plantlets.     

Development of a regeneration protocol through indirect organogenesis in prickly pear cactus (Opuntia ficus-indica (L.) Mill)

An indirect organogenesis regeneration protocol for Opuntia ficus-indica (L.) Mill var “Blanco sin Espinas” is described. One centimeter square cladode explants sections from previously micropropagated prickly pear plants were cultured in Murashige and Skoog (MS) basal medium supplemented with 20 different combinations of 2,4-dichlorophenoxy acetic acid (2,4-D) and benzyladenine (BA). The best calli induction and regeneration response were observed when 2.26 μM 2,4-D and 2.21 μM BA combination was applied to the nopal explants. Regenerating calli was capable of forming new buds when transferred to MS basal medium supplemented with 0.5 μM BA (proliferation medium). Shoot elongation and rooting were achieved on MS medium without plant growth regulators. Excellent acclimatization to greenhouse conditions was observed for all transferred plantlets. By this procedure no morphological differences were observed between the regenerated and mother plants. This protocol may be also utilized to carry out plant regeneration after genetic transformation, in order to develop transformed plants without the presence of chimeric zones.     

Dry-matter partitioning in a tomato crop: Comparison of two simulation models

SUMMARY

TOMSIM(l.O) and TOMGRO(I.O) are two dynamic models for tomato growth and development. Their sub-models for dry-matter distribution between leaves, stem and fruits were compared and discussed. In both models the simulated dry-matter distribution is regulated by the relative sink strengths of the plant organs. These sink strengths are quantified by the potential growth rates of individual organs, i.e. the growth rates under conditions of non-limiting assimilate supply. This approach is general and not limited to the tomato crop. In TOMGRO(J.O), fruits, leaves and internodes stay within age classes and move from class to class during development, whereas in TOMSIM (1.0), record is kept of every fruit truss separately but leaves and internodes are lumped together (i.e. no record of weight or leaf area per age class as in TOMGRO(l.O)). In TOMSIM(1.0), vegetative sink strength is a constant, whereas in TOMGRO(l.0) it is calculated from potential area expansion rate of leaves and specific leaf area. In both models, the ratio between leaf growth and stem growth is constant. In TOMGRO(l.O) there is a feed-back mechanism which controls the vegetative/generative balance: a higher demand/supply ratio for assimilates induces higher fruit abortion rates. In TOMSIM(l.O) the number of fruits set per truss is not simulated, but is an input to the model. TOM SIM (1.0) functions representing flowering rate, fruit growth period, vegetative sink strength and fruit sink strength were compared with similar TOMGRO(l.O) functions, in their dependence on temperature and physiological plant age. A sensitivity analysis was made for the effects of temperature, flowering rate, and fruit and vegetative sink strengths on dry-matter distribution for both models. A validation of both models was based upon periodic destructive harvests in: 1) a greenhouse experiment in Wageningen, using a round tomato cultivar, consisting of a control treatment and a treatment where every second truss was removed at anthesis, and 2) two greenhouse experiments conducted in Montfavet, using a beefsteak tomato cultivar. Daily shoot dry-weight increase, average 24 h greenhouse temperatures and numbers of fruits set per truss (in TOMGRO(l.O) numbers of flowers per truss) were inputs to the models. In general dry-matter distribution was simulated well by both models for the cultivar and conditions where they were developed. TOMGRO(1.0)'s poor performance in one of the validations resulted from the absence of an assimilate storage pool. To achieve reasonable agreement between measurements and simulations for situations other than where the models were developed, parameter adjustments had to be made, most likely reflecting cultivar differences. Strong and weak points of both models are discussed.     

Dry-matter production in a tomato crop: comparison of two simulation models

TOMSIM(l.O) and TOMGRO(l.O) are two dynamic models for tomato growth and development. Their submodels for dry matter production are compared and discussed. In TOMSIM(l.O), dry matter production is simulated by a modified version of SUCROS87 (Spitters et al., 1989). Single leaf photosynthesis rates are calculated separately for shaded and sunlit leaf area at different depths in the canopy, according to the direct and diffuse components of light; daily crop gross assimilation rate (A) is computed by integration of these rates over the different depths and over the day. In TOMSIM(l.O) leaf photochemical efficiency (e) and potential leaf gross photosynthesis rate at saturating light level (Pgmax) both depend on temperature and C02 level. In TOMGRO(l.O) crop gross photosynthesis rate is calculated by the equation of Acock et al. (1978); e is a constant and Pg max is a linear function of C02. In both models leaf photosynthesis characteristics are assumed to be identical in the whole canopy. Maintenance respiration (Rm) and conversion efficiency (Cf) are taken into account in the same way, except that root maintenance respiration is neglected in TOMGRO(l.O). For both models a sensitivity analysis was performed on the input variables (light intensity, temperature, C02 and leaf area index (LAI)) and on some of the model parameters. Under most conditions considered, simulated A was found to be 5-30% higher in TOMSIM(l.O) than in TOMGRO(l.O). At temperatures above 18°C Rm was also higher in TOMSIM(l.O), and C, was 4% higher in TOMGRO(l.O). The two models were very sensitive to changes in e and to a lesser extent to changes in the light extinction coefficient, whereas the scattering coefficient of leaves had hardly any effect on the simulated A. TOMGRO(l.O) appeared to be rather sensitive to the C02 use efficiency, whereas at ambient C02 level mesophyll resistance was quite important in TOMSIM(l.O). Four sets of experimental data (differences in cultivar, C02 enrichment and planting date) from Wageningen (The Netherlands) and Montfavet (southern France) were used to validate the models. Average 24 h temperature and average daily C02 concentration values were used as input to the models. For the Wageningen experiments, hourly PAR values were calculated from the daily global radiation sum by TOMSIM(l.O) and used as input in both models. For the Montfavet experiment, average hourly PAR measurements were used. Also measured LAI, dry matter distribution and organ dry weights (for calculation of Rm) were input to the simulation. In the Wageningen experiments, total dry matter production was simulated reasonably well by both models, whereas in the Montfavet experiment an under-estimation of about 35% occurred. TOMGRO(l.O) and TOMSIM(l.O) simulated almost identical curves in all four experiments. Strong and weak points of both models are discussed.     

Studies on the post-initiation development of flower buds of tomato (Lycopersicon Esculentum)

The application of high temperature (ca. 70° F., 21° C.) to tomato plants for varying periods prior to first anthesis showed that:

1. Inflorescence abortion occurred when high temperature (and low light) was applied in the late stages of development of the inflorescence. Similar treatment in the early stages, up to the time when the first cluster of buds could be seen by the naked eye, did not induce abortion.

2. In low light conditions, equivalent to those occurring naturally at mid-winter, an initial period of high temperature (70° F.) followed by low (60° F., 15–5° C.) induced a greater flowering capacity than constant low temperature.

3. The beneficial effects on flower development resulting from early high temperature treatment were greatest when the day temperature was high, though some benefit also resulted from a high night temperature. The effects were evident only with early (October) sown plants. There was no apparent adverse effect on flower development of high temperature applied up to the “visible bud” stage (2,250 kilolux hours).     

8年以上温室冬春茬番茄平衡施肥技术研究

为了改变生产者在保护地蔬菜种植上施用化肥的盲目性、随意性和轻视有机肥的现象,解决因保护地土壤退化、养分比例失调而造成的土壤养分供应障碍,生理性病害普遍发生的问题,采用大田试验方法,于2005-2007年在连作番茄8年以上温室内,开展了生产上普遍应用的习惯施肥与全部用化肥、化肥+有机肥平衡施肥3种施肥方法对番茄的影响对比试验研究。结果表明,2种平衡施肥处理较习惯施肥,可显著改善植株生物学性状,减轻番茄生理性病害,改进番茄品质;极显著地提高番茄产量和效益,其中667m2分别增产27.5%和34.9%,分别增收4413.7和4872.1元;平衡施肥以化肥+有机肥在培肥地力,改善番茄品质,提高产量和经济、社会、生态等综合效益上更优更符合农业发展方向,值得在生产上推广应用。     

Development of a transformation protocol for Tecomella undulata (Smith) Seem from cotyledonary node explants

A protocol was optimized for genetic transformation of Rohida (Tecomella undulata) from cotyledonary node tissues using Agrobacterium tumefaciens strain GV2260 harboring binary vector pBinAR containing osmotin and nptII gene under control of CaMV35S promoter. This is the first report on the transformation of T. undulata. The effective concentration of selectable marker antibiotic used for screening of transformants was 85.97 μM in shooting media and 42.98 μM in rooting media. PCR and Southern blot confirmed integration of osmotin gene into genome of Rohida.     

Nutritional control of a nutrient-film tomato crop

The hypothesis that adequate control of the nutrient supply of an NFT tomato crop could be obtained merely by measuring the pH and electrical conductivity of the nutrient solution, was tested. Data are presented from 27 weekly chemical analyses of the nutrient solution. The crop yield was good and the results suggested (a) that pH and electrical conductivity measurements provide adequate control when used in conjunction with observations on plant appearance; (b) that even further simplification of nutritional management could be achieved given further development work.     

Influence of sub-optimal temperature on tomato growth and yield: a review

Summary

The effects of temperature on growth, development and yield of tomato (Lycopersicon esculentum) are reviewed with special emphasis on cultivar differences. The focus is on sub-optimal temperatures, above the level where chilling injury occurs. Temperature has a large effect on all aspects of development. Leaf and truss initiation rates decrease linearly with decreasing temperature. Although these rates may be different for different cultivars their response to temperature is the same. Young plants grown at sub-optimal temperatures produce thicker leaves, so they intercept less light and therefore have a lower relative growth rate. There was no interaction between temperature and cultivar for relative growth rate and related traits. In a crop producing fruits, this aspect is less important as most of the light is intercepted anyway. At sub-optimal temperatures, fruit set is reduced as a result of poorer pollen quality. The period between anthesis and ripening of the fruit increases and, as the growth rate of the fruit at a certain developmental stage is independent of temperature, fruits become larger at sub-optimal temperature. Higher temperature leads to an increase in early yield, at the cost of vegetative growth, but may also cause a delay in later trusses. Total yield over a whole season might be equal at lower temperatures, but higher tomato prices early in the season do not make it economically profitable to reduce the temperature in the greenhouse. Short-term effects might thus be different from long-term effects. In the literature, the link between yield and whole plant growth is often missing, limiting the possibilities of studying the underlying processes that contribute to changes in yield. Breeding for cultivars with equal production at lower temperatures is hampered by the limited variation for temperature response in cultivated tomato. Therefore breeders have to look for other sources of variation, as in related wild Lycopersicon species.     

Protocol: An improved high-throughput method for generating tissue samples in 96-well format for plant genotyping (Ice-Cap 2.0)

Background  

We previously developed a high-throughput system called 'Ice-Cap' for growing Arabidopsis seedlings in a 96-well format and rapidly collecting tissue for subsequent DNA extraction and genotyping. While the originally described Ice-Cap method is an effective tool for high-throughput genotyping, one shortcoming of the first version of Ice-Cap is that optimal seedling growth is highly dependent on specific environmental conditions. Here we describe several technical improvements to the Ice-Cap method that make it much more robust and provide a detailed protocol for implementing the method.     

Effect of diurnal fluctuations in humidity on the accumulation of nutrients in the leaves of tomato (Lycopersicon esculentum)

Tomato plants were grown at four humidities (vapour pressure deficits of 0.15,0.25,0.43 and 0.65 kPa) that were maintained continuously for four weeks. The Ca and Mg contents of the upper leaves declined progressively with increasing humidity. In the main experiment, a high (0.21 kPa) and low humidity (0.47-0.55 kPa) during the day were tested in factorial combination with a high (0.16 kPa) and low humidity (0.45-0.50 kPa) at night. Leaves were divided into terminal and basal leaflets, the rest of the lamina and the petiole. High humidity at night reduced the Ca, Mg, P and K contents (%) of all portions of the leaf lamina whilst high humidity during the day reduced only the Ca and K contents. The N content was not affected by humidity. The Ca and K contents of the petioles were reduced by high humidity during the day and at night. The changes in the Ca, Mg and K contents of the tissue in response to high humidity were greater in the terminal leaflet than in the basal leaflet or in the rest of the lamina, suggesting that the terminal leaflet was suitable for monitoring the nutrient status of the crop. Yellowing of the terminal leaflet at high humidity appeared to be associated with deficiencies of both Ca and K rather than with Ca only.     

Differential utilization of nitrates by solanaceous species,crops (tomato and pepper) and weeds (black nightshade and thorn apple)

Summary

The capacity of nitrate absorption, accumulation and reduction was studied in four species of the Solanaceae. Two were cultivated species, tomato (Lycopersicon esculentum Mill.) and pepper (Capsicum annuum L.), and two weed species, black nightshade (Solanum nigrum L.) and thorn apple (Datura ferox L.), which often infest these crops. Black nightshade and thorn apple absorbed nitrates most rapidly. Maximum nitrate accumulation was observed earlier in tomato than in the other species, and as in pepper coincided with the flower bud stage. Maximum nitrate accumulation in black nightshade and thorn apple took place during flowering. Only in tomato and thorn apple did maximum nitrate reductase activity occur at the time of maximum nitrate accumulation. Growth and soil nitrate absorption was slowest in pepper, but nitrate accumulation by the plant and nitrate reductase activity was more sustained than in the other species. It was concluded that if these weeds emerge simultaneously with the crops, competition for nitrates by thorn apple, and especially black nightshade, would be greater than for the tomato crop and even more so for the pepper crop.     

Genetics of resistant to tomato yellow leaf curl virus in tomato

The genetics of resistance to tomato yellow leaf curl virus (TYLCV) were studied in TYLCV-resistant lines developed by crossing wild species of Lycopersicon pimpinellifolium, L. hirsutum and L. peruvianum resistant to TYLCV with susceptible L. esculentum cv. Special Back. Crosses between TYLCV-resistant lines derived from the same wild species produced progenies similar to their parents in their level of resistance. However, progenies from interspecific crosses showed greater resistance than either parent suggesting that the genes for TYLCV resistance contributed by different wild species are probably not the same (non-allelic). The gene action for TYLCV resistance also varied with the source of resistance. Analysis of F₁, F₂ and BC populations for lines derived from L. pimpinellifolium showed that resistance to TYLCV in these lines is a quantitative trait with some dominance.     

Role of plant age in the resistance of Lycopersicon hirsutum f. glabratum to the tomato leafminer Tuta absoluta (Lepidoptera: Gelechiidae)

The objective of this work was to study the role of plant age in the resistance of tomato, Lycopersicon hirsutum, to the tomato leafminer Tuta absoluta (Meyrick). Determination of the levels of tridecan-2-one and undecan-2-one in L. hirsutum were made at three different ages (2–4 months after germination), as well as the leaf area, and the density and types of trichomes present on L. hirsutum and L. esculentum. This information gathered was correlated with the following biological characteristics of T. absoluta: rates of oviposition and egg eclosion; length of egg, larval and pupal stages; mortality in the larval and pupal stages; pupal weight and proportion of females. T. absoluta showed higher oviposition and egg eclosion, as well as lower mortalities and shorter larval and pupal periods in L. esculentum than in L. hirsutum. The proportion of females obtained was also higher on L. esculentum. The rate of egg eclosion and proportion of females as well as the length of the larval stage and the mortality of larvae appeared to be affected by plant age. A higher proportion of females, mortality of larvae and length of the larval period were obtained with older plants of L. hirsutum, while the rate of egg eclosion was higher for L. hirsutum plants that were 3 months old. For L. esculentum, plant ageing increased larval mortality. It seems that the glandular trichome density in L. hirsutum increased with plant age leading to an increase in the levels of tridecan-2-one, which slows larval development.     

Comparative effects of regulated deficit irrigation (RDI) and partial root-zone drying (PRD) on growth and cell wall peroxidase activity in tomato fruits

The effects of regulated deficit irrigation (RDI) and partial root-zone drying (PRD) on tomato fruit growth and cell wall peroxidase activity in tomato exocarp were investigated in growth chamber conditions. The RDI treatment was 50% of water given to fully irrigated (FI) plants and the PRD treatment was 50% of water of FI plants applied to one half of the root system while the other half dried down, with irrigation shifted when soil water content of the dry side decreased 15–20%. RDI significantly reduced fruit diameter, though PRD reduced fresh weight while having no significant effect on fruit diameter. The activity of peroxidase was significantly higher in RDI and PRD treated plants compared to those of FI. Differences between RDI and PRD were expressed on temporal basis. In the fruits of RDI treated plants peroxidase activity began to increase in the phase when fruit growth started to decline with the peak of enzyme activity of 6.1 HRPEU g⁻¹ FW reached in the phase of mature green fruits when fruit growth rate was minimal. Increase of peroxidase activity in PRD fruits coincided with the ripening phase and the peak of enzyme activity (5.3 HRPEU g⁻¹ FW) was measured at the end of fruit ripening. These data potentially identified contrasting and different roles of tomato exocarp cell wall peroxidase in RDI and PRD treated plants. In RDI treated plants peroxidase may have a role in restricting fruit growth rate, although the increase in enzyme activity during ripening of PRD treated fruit pointed out that peroxidase may also control fruit maturation by inducing more rapid process.     

Improved protocol for Agrobacterium mediated transformation of tomato and production of transgenic plants containing carotenoid biosynthetic gene CsZCD

Improved protocol for Agrobacterium mediated transformation of tomato (Lycopersicon esculentum) Micro-Tom was developed to use in corporation of the carotenoid biosynthetic genes CsZCD (Crocus zeaxanthin 7,8-cleavage dioxygenase). From these experiment, a transformation methodology using explants from cotyledons cultured for 1 day on the medium with zeatin 2 mg/L, IAA 0.1 mg/L, carefully submerged in the Agrobacterium inoculum for 20 min, then concultured with the agrobacterium for 3 days on the same medium, followed by a transfer to the same medium with 500 mg/L cefotaxin for 3 days and then by a transfer to the same medium with 100 mg/L kanamycin and 500 mg/L carabenillin for 6–8 weeks and resulted in a greater than 20% transformation efficiency in the concentration of Agrobacterium OD600 = 0.2 tested. In this transformation method, no feeder layers were used and the subculture media was minimal. Among the Agrobacterium concentrations of OD600 = 0.2, 0.5 and 1.0, the best transformation efficiency, 20.87%, was obtained by using OD600 = 0.2, which was significantly higher than that of OD600 = 1.0. The presence of the inserted target genes was checked using a rapid and efficient PCR test. The protocol was successfully employed in the production of transgenic Micro-Tom tomato containing the carotenoid biosynthesis CsZCD under constructive promoter. This procedure represents a simple, efficient and general means of transforming tomato.     

利用多重PCR反应同时鉴定番茄Ty-3a基因和抗溃疡病QTL

番茄黄化曲叶病毒病和番茄溃疡病是两种分布广泛、危害严重的世界性病害,严重影响了番茄的产量,是国内外番茄抗病育种的重要目标。为了提高抗病育种的效率,研究建立了抗番茄黄化曲叶病毒病Ty-3a基因和抗番茄溃疡病QTL的多重PCR反应体系。该体系利用同一PCR反应,对分别与番茄抗黄化曲叶病毒病的Ty-3a基因和抗番茄溃疡病的一个QTL位点Rcm5.1紧密连锁的标记进行了扩增、筛选,扩增的特异性片段与单引物扩增片段吻合。其中与Ty-3a基因紧密连锁的标记Ty-3F/Ty-3R为共显性标记,基因型为Ty-3a/Ty-3a的材料均产生630 bp的特异性片段,基因型为ty-3/ty-3的材料均产生320 bp的特异性条带,基因型为Ty-3a/ty-3的材料产生630 bp和320 bp的特异性片段;与番茄溃疡病QTL位点Rcm5.1连锁的标记TG318F/TG318R为显性标记,只有抗病材料产生580 bp的特异性片段,感病材料不产生特异性条带。经反复验证结果准确、稳定,可用于同一PCR反应体系中对番茄抗黄化曲叶病毒病Ty-3a基因和番茄溃疡病QTL的同时筛选鉴定。