A temporary immersion plant propagation bioreactor with decoupled gas and liquid flows for enhanced control of gas phase

Temporary immersion bioreactors (TIBs) are being used to propagate superior plant species on a commercial scale. We demonstrate a new TIB design, a Hydrostatic‐driven TIB (Hy‐TIB), where periodic raising and lowering the media reservoir maintains the advantages of temporary immersion of plant tissues without requiring large amounts of gas to move the media that is a characteristic of other TIB designs. The advantage of utilizing low volumes of gas mixtures (that are more expensive than air) is shown by a doubling of the growth rate of plant root cultures under elevated (40%) oxygen in air, and with CO₂ supplementation showing improved phototrophic and photomixotrophic growth of seedless watermelon meristem cultures. The development of this bioreactor system involved overcoming contamination issues associated with utilizing very low gas flow rates and included utilizing microchip pressure sensors to diagnose unexpected changes in internal bioreactor pressure (± 20 Pa ～0.0002 atm) caused by flexing of non‐rigid plastic bag vessels. The overall design seeks to achieve versatility, scalability and minimum cost such that bioreactor technology can play an increasing role in the critical need to improve plant productivity in the face of increasing demand for food, reduced resources, and environmental degradation. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:337–345, 2016     

Effects of photo-oxidative loss of FeNa2EDTA and of higher iron supply on chlorophyll content, growth and propagation rate in triploid watermelon cultures

Summary Triploid watermelon (Citrullus lanatus [Thunb.] Matsum. & Nakai) Arka Manik cultures exhibited a general pale color and yellowing of lower leaves by 3–4 wk following subculturing to fresh medium. Protecting the medium from light by covering the surface with sterile aluminum foil significantly improved leaf color and chlorophyll content by 50–60%. Raising the concentration of FeNa₂EDTA (Fe-EDTA) in the light-exposed medium from the normal level (100 μM) to 150 or 200 μM significantly increased the leaf chlorophyll two- to threefold, suggesting that the light-induced effect was mainly attributable to photo-oxidation of Fe-EDTA. Fe-EDTA supplementation also resulted in better expansion and lobe formation of upper leaves, a reduction in internode length, an increase in the number of nodes, and a significant improvement in propagation rate. There was also a reduction in axillary sprout growth at elevated Fe-EDTA levels, indicating a more prominent apical dominance effect and a reduction in the number and vigor of tendrils. Increasing the Fe-EDTA level in the medium to 150 μM is suggested for routine micropropagation of triploid watermelon, in order to compensate for the photo-oxidative loss of Fe-EDTA.     

种子小蜂和空壳果实对生物能源树种黄连木果实和油产量的影响(英文)

黄连木Pistacia chinensis Bunge因果实含油量高, 被作为生物能源树种在我国进行大面积栽培推广和能源开发利用。然而, 专性寄生害虫（主要为黄连木广肩小蜂Eurytoma plotnikovi Nikolskaya）的危害及由单性结实和败育所造成的空壳果实可能影响黄连木的果实产量和质量。为了评价种子害虫和空壳果实对黄连木果实产量和质量的影响, 我们测定了河南省济源市3个黄连木种群40株结果雌树的果实产量和油产量, 同时测定了完好果实、 空壳果实和虫蛀果实等3类果实的干重和含油量。结果表明：（1）虫害率和空壳率分别达22.6%和53.0%, 且不同个体和种群间均存在显著差异（P<0.05）。（2）空壳果实（30.7 mg/果）和虫蛀果实（33.1 mg/果）的干重均不到完好果实（67.4 mg/果）的一半; 空壳果实（3.9%）和虫蛀果实（3.8%）的含油量均显著低于完好果实（39.9%）。（3）实测的果实产量（2.9 kg/树）和油产量（0.6 kg/树）分别仅为期望产量的50%（4.7 kg/树）和33%（1.9 kg/树）, 但在3个种群之间无显著差异。本研究显示小蜂危害和空壳果实对黄连木果实产量和质量能造成严重影响, 在将其作为生物能源树种利用时对此应加以克服。     

Using urea phosphate to enhance the effect of gibberellin A3 on grape size

Gibberellic acid (GA₃) is widely used to enlarge the berries of seedless grapes (Vitis vinifera L). In cv. Sultana (Thompson Seedless) the addition of 1000 mg/L urea phosphate (UP) to GA₃ solutions after fruit set reduced the pH of the solutions to a stable pH 2.9 and enhanced the effect of GA₃ on berry size and delayed maturation. Addition of citrate buffer, pH 2.9, to GA₃ sprays did not affect berry size or maturation. The possibility of improved GA penetration due to the low pH is considered. The nutritional effect of UP and direct enhanced penetration by the urea ion are also discussed.Contribution from the Agricultural Research Organization, The Volcani Center, Bet Dagan, Israel. No. 1735-E, 1986 series.     

Use of fluorescein diacetate (FDA) to determine ploidy of in vitro watermelon shoots

Ploidy of watermelon [Citrullus lanatus (Thunb.) Matsum. and Nakai shoots and plantlets was estimated by painting the lower epidermis of intact in vitro-derived leaves with fluorescein diacetate (FDA) and observing fluorescence of guard cell chloroplasts with a microscope and UV light. Leaves from in vitro shoot-tip cultures of known diploid cultivars and tetraploid breeding lines were used to establish the mean number of chloroplasts per guard cell pair. Leaves from diploid and tetraploid shoot cultures had 9.7 and 17.8 chloroplasts per guard cell pair, respectively. This method then was used to estimate the ploidy of shoots regenerated from cotyledon explants of the diploid cultivar Minilee. Approximately 11% of the 188 regenerated shoots were classified as tetraploid during in vitro culture. Putative tetraploids were transplanted to the field and self-pollinated. About 45% of tetraploids identified in vitro produced fruit and viable seed. Chloroplast counts of R₁ progeny were used to confirm their ploidy. All of the putative diploids were confirmed diploid and all putative tetraploids proved to be non-chimeric true breeding tetraploids. This revised version was published online in June 2006 with corrections to the Cover Date.     

Cleansing the long-term micropropagated triploid watermelon cultures from covert bacteria and field testing the plants for clonal fidelity and fertility during the 7–10 year period in vitro

The study was aimed primarily at cleansing the in vitro-decline displaying long-term micropropagated triploid watermelon ‘Arka Manik’ cultures from covert bacteria and their further field testing. Disinfectant treated shoots showed endophytic survival but to a lesser extent in shoot-tips. Culturing the NaOCl (5 min) treated shoot-tips on filter paper bridges in liquid watermelon medium containing single antibiotic (gentamycin (Gm), amoxycillin (Ax) or cefazolin (Cz) at 50, 250, 500 or 1000 mg l⁻¹) for 1 month followed by repeated indexing of medium and tissue for two-four subculture passages facilitated the cleansing of cultures with 12.5% recovery as monitored for 2 years. Partial bleaching damage by NaOCl, phytotoxicity to higher levels of antibiotics, poor growth response in the initial sucrose-free medium and rampant hyperhydricity came in the way of a higher recovery. The effectiveness of the above approach was ascertained after back inoculating clean cultures with a mixture of Gram-positive and Gram-negative bacteria yielding 30, 45 and 35% clean cultures in Gm, Ax and Cz (250 mg l⁻¹ for 1 month) treatments, respectively in modified medium compared with 10% recovery after mere HgCl₂ surface sterilization. The results indicated that antibiotic treatment was essential but not its choice, and extended culture-indexing subsequent to disinfection or antibiotic treatment was crucial in identifying clean stocks. Cleansed cultures, showed restoration of growth but a drop in rooting. Most of the in vitro cultures appeared normal and true-to-type during the 7–10 year period in vitro but a small proportion of bacteria-harboring stocks displayed ‘epigenetic variations’. Acclimatized plants and those in the field also appeared true-to-type but for a minor proportion derived from bacteria-harboring stocks. Field-plants which originated from bacteria-freed stocks after 9 years of continuous culturing were normal and fertile validating the possibility of keeping treasured cultures in vitro for long periods if covert contaminants are checked.     

无籽西瓜品种SSR指纹图谱构建及遗传多样性分析

利用SSR标记构建了27份中国无籽西瓜主栽品种的DNA指纹并进行了遗传多样性分析。24对多态性引物共扩增出66种基因型,基因型数2-5个不等,平均2.75个。平均多态性信息量(PIC)为0.37,变化范围为 0.19～0.66。有4个品种具有特征谱带。27个品种遗传相似系数变化范围为 0.7045～1.0,平均0.8683。组合24对引物,除无法区分‘郑抗无籽1号’与‘雪峰花皮无籽’,其余品种均能一一区分开。采用类平均法进行聚类分析,在相似系数0.83处,可将27个品种分为3大类。     

Identification and characterisation of the tomato parthenocarpic mutant high fruit set under stress (hfs) exhibiting high productivity under heat and salt stress

Pollination and therefore fruit set in tomato (Solanum lycopersicum) is very sensitive to temperature. Parthenocarpy can be a very useful trait in tomato breeding in sustainable agriculture faced with global warming. Within a collection of Moneymaker tomato mutants a monogenic and recessive mutant with high fruit number was identified in heat stress conditions and named high fruit set under stress (hfs). No morphological alterations in vegetative and reproductive organs were observed except the bigger size of ovary. hfs is parthenocarpic, pollination not being required for fruit set, although this trait was not absolute since it produced some under‐seeded fruit. When plants were grown under extreme temperatures (higher than 35°C), hfs exhibits higher fruit yield than wild‐type (WT) due to increased fruit number. Another very interesting characteristic of hfs is its improved fruit quality under heat stress, exhibiting a better sweetness/acidity balance than WT. Interestingly, hfs was also tolerant to the combination of heat and salt stress, and the positive effect on production was due to both components of yield, fruit number and fruit weight. The generation of mostly seedless fruit and the high productivity and fruit quality under extreme temperatures make hfs a very interesting mutant to obtain new breeding high‐yield lines in adverse environmental conditions.     

葡萄VvRGL基因应答GA调控无核果实发育的研究

DELLA蛋白是GA信号传导途径的核心作用元件,属于GRAS核转录调节因子家族。该研究以‘魏可’葡萄品种为试材,通过基因克隆、启动子分析、染色体定位、基因蛋白结构及系统进化分析,鉴定VvRGL1和VvRGL2精确序列,预测其潜在功能,并采用qRT-PCR技术检测VvRGL基因应答GA在果皮、果肉及种子(区)的时空表达特征。结果表明:(1)VvRGL1及VvRGL2的染色体定位为Chr14和Chr7,开放阅读框(ORF)为2 007bp和1 815bp,编码氨基酸数量为668和604个;二者均含有GRAS保守结构域,但却不具备DELLA结构域,属于GRAS转录因子大家族成员,而不属于DELLA亚家族;基因结构分析表明,VvRGL1的DNA序列有1个内含子,2个外显子,而VvRGL2的DNA序列无内含子,有1个外显子,基因结构高度保守;进化分析表明,VvRGL1与拟南芥和柑桔的亲缘关系较近,而VvRGL2与草莓和杨树的亲缘关系较近。(2)2个基因的启动子均含有响应赤霉素和胚乳发育相关的作用元件,表明它们可能参与响应GA信号传导和种子胚乳发育过程。(3)qRT-PCR结果显示,外源GA处理均不同程度降低了2个基因在葡萄果皮和种子区的表达,但却上调了其在幼果期果肉中的表达,表明在果皮与种子(区)中,GA可能通过抑制葡萄VvRGL1/RGL2基因的表达参与调控葡萄无核果实的发育。本研究结果为进一步阐明VvRGL在赤霉素信号传导及葡萄无核果实发育机理中的作用机制提供重要依据。