Daniel J. Cantliffe

Melon Fruits: Genetic Diversity, Physiology, and Biotechnology Features

Among Cucurbitaceae, Cucumis melo is one of the most important cultivated cucurbits. They are grown primarily for their fruit, which generally have a sweet aromatic flavor, with great diversity and size (50 g to 15 kg), flesh color (orange, green, white, and pink), rind color (green, yellow, white, orange, red, and gray), form (round, flat, and elongated), and dimension (4 to 200 cm). C. melo can be broken down into seven distinct types based on the previously discussed variations in the species. The melon fruits can be either climacteric or nonclimacteric, and as such, fruit can adhere to the stem or have an abscission layer where they will fall from the plant naturally at maturity. Traditional plant breeding of melons has been done for 100 years wherein plants were primarily developed as open-pollinated cultivars. More recently, in the past 30 years, melon improvement has been done by more traditional hybridization techniques. An improvement in germplasm is relatively slow and is limited by a restricted gene pool. Strong sexual incompatibility at the interspecific and intergeneric levels has restricted rapid development of new cultivars with high levels of disease resistance, insect resistance, flavor, and sweetness. In order to increase the rate and diversity of new traits in melon it would be advantageous to introduce new genes needed to enhance both melon productivity and melon fruit quality. This requires plant tissue and plant transformation techniques to introduce new or foreign genes into C. melo germplasm. In order to achieve a successful commercial application from biotechnology, a competent plant regeneration system of in vitro cultures for melon is required. More than 40 in vitro melon regeneration programs have been reported; however, regeneration of the various melon types has been highly variable and in some cases impossible. The reasons for this are still unknown, but this plays a heavy negative role on trying to use plant transformation technology to improve melon germplasm. In vitro manipulation of melon is difficult; genotypic responses to the culture method (i.e., organogenesis, somatic embryogenesis, etc.) as well as conditions for environmental and hormonal requirements for plant growth and regeneration continue to be poorly understood for developing simple in vitro procedures to culture and transform all C. melo genotypes. In many cases, this has to be done on an individual line basis. The present paper describes the various research findings related to successful approaches to plant regeneration and transgenic transformation of C. melo. It also describes potential improvement of melon to improve fruit quality characteristics and postharvest handling. Despite more than 140 transgenic melon field trials in the United States in 1996, there are still no commercial transgenic melon cultivars on the market. This may be a combination of technical or performance factors, intellectual property rights concerns, and, most likely, a lack of public acceptance. Regardless, the future for improvement of melon germplasm is bright when considering the knowledge base for both techniques and gene pools potentially useable for melon improvement.

Somatic embryogenesis and plant regeneration in tissue cultures of sweet potato (Ipomea batatas Poir.)

Leaf, shoot-tip, stem, and root explants of sweet potato (Ipomea batatas Poir.) gave rise to two kinds of callus on nutrient agar medium containing 0.5 to 2.0 mg/l 2,4-D. One callus, bright- to pale-yellow, was compact and organized, while the other was dull-yellow and friable. The former callus gave rise to numerous globular and heart-shaped embryoids. When transferred onto hormone-free medium, the embryoids readily developed into a torpedo-shape before germination. The plantlets were transplanted to soil where they flowered and formed storage roots at maturity.

Transgenic sweet potato plants obtained byAgrobacterium tumefaciens-mediated transformation

Stable expression of foreign genes was achieved in sweet potato (Ipomoea batatas (L.) Lam) plants using anAgrobacterium tumefaciens mediated system. Embryogenic calluses produced from apical meristems of cultivar White Star were multiplied and cocultivated withA. tumefaciens strain EHA101 harboring a binary vector containing the β-glucuronidase (GUS) and neomycin phosphotransferase (NPT II) genes. The calluses were transferred to selective regeneration medium and kanamycin resistant embryos were recovered which developed into morphologically normal plants. Histochemical and fluorimetric GUS assays of plants developed from the kanamycin resistant embryos were positive. Amplified DNA fragments were produced in polymerase chain reactions using GUS-specific primers and DNA from these plants. Transformation was confirmed by Southern analysis of the GUS gene. With the developed method, transgenic sweet potato plants were obtained within 7 weeks. This method will allow genetic improvement of this crop by the introduction of agronomically important genes.

Somatic Embryogenesis and the Technology of Synthetic Seed

Synthetic seeds are functionally defined as somatic embryos engineered to be of use in commercial plant production (Gray 1990b). The actual form of synthetic seed (i.e., presence or absence of a synthetic seed coat, whether they are hydrated or dehydrated, quiescent or not, etc.) may vary depending on the specific crop application.

Selective enhancement of Ipomoea batatas Poir. embryogenic and non-embryogenic callus growth and production of embryos in liquid culture

Embryogenic callus cultures of Ipomoea batatas Poir. produce fast growing non-embryogenic material which soon dominates the cultures. Our objective was to selectively enhance the proliferation of the embryogenic fraction. For this, the effect of BAP and 2,4-D concentrations on growth of embryogenic and non-embryogenic callus were studied and consequently, nutrient media for the production and indefinite maintenance of embryogenic callus without embryo formation were defined. Selective proliferation of embryogenic callus was obtained on solid media with 10 μM 2,4-D and 1 μM BAP and in liquid media with 5 μM 2,4-D. Selective proliferation of non-embryogenic callus occurred in liquid medium with 1 μM 2,4-D. In embryogenic liquid culture, embryos were produced with 0–2 μM 2,4-D. Increasing 2,4-D concentration from 0 to 2 μM in these cultures restricted embryo development.

Stable transformation of lettuce cultivar South Bay from cotyledon explants

Transgenic plants of lettuce cultivar (cv.) ‘South Bay’ were produced by using Agrobacterium tumefaciens vectors containing the β-glucuronidase (GUS) reporter gene and the NPT II gene for kanamycin resistance as a selectable marker. High frequency of transformation, based on kanamycin resistance and assays for GUS expression, was obtained with 24 to 72-h-old cotyledon explants cocultivated for 48 h with Agrobacterium tumefaciens. After the cocultivation period, the explants were placed in selection medium containing 50 or 100 mg l−1 of kanamycin, 100 mg l−1 cefotaxime and 500 mg l−1 carbenicillin for 10 days. Surviving explants were transferred every 14 days on shoot elongation medium. Progenies of R0 plants demonstrated linked monogenic segregation for kanamycin resistance and GUS activity.

Ethylene evolution and endo-beta-mannanase activity during lettuce seed germination at high temperature

Altas temperaturas durante a embebicao das sementes de alface podem atrasar ou inibir a germinacao e o endosperma parece ser o responsavel na restricao da protrusao da radicula. O envolvimento da enzima endo-b-mananase durante a germinacao de sementes de alface a 35°C e a influencia do etileno na regulagem desta enzima foram estudados. Sementes das cultivares Dark Green Boston (DGB) e Everglades (EVE) foram germinadas em agua ou em solucoes de 10 mmol L-1 de 1-aminociclopropano-1-acido carboxilico (ACC), 10 mmol L-1 de amino-etoxi-vinil-glicina (AVG), ou 20 mmol L-1 de tiossulfato de prata (STS). Sementes foram ainda osmoticamente condicionadas em solucoes de polietilenoglicol (PEG), ou PEG + ACC, PEG + AVG, ou PEG + STS. Sementes nao tratadas germinaram 100% a 20°C. A 35°C, EVE germinou 100%, enquanto DGB germinou somente 33%. A germinacao a 35°C aumentou em sementes osmoticamente condicionadas ou sementes que receberam ACC durante a incubacao. Maior evolucao de etileno foi detectada em EVE do que em DGB durante a germinacao a 35°C. AVG nao inibiu a germinacao de DGB a 35°C, enquanto que STS inibiu. Maior atividade de endo-b-mananase nas sementes foi observada em EVE quando comparado com DGB. Fornecimento de ACC tanto durante o condicionamento osmotico como durante a germinacao, aumentou a atividade de endo-b-mananase, enquanto que AVG e STS proporcionaram um decrescimo ou ausencia da atividade enzimatica. O etileno pode minimizar o efeito inibitorio de altas temperaturas na germinacao de sementes de genotipos sensiveis de alface devido ao aumento da enzima endo-b-mananase, possivelmente levando ao enfraquecimento do endosperma.

Somatic embryony patterns and plant regeneration inIpomoea batatas poir

SummaryPatterns of somatic embryo development from callus were elucidated inIpomoea batatas Poir. (sweet potato). Embryos at three stages of development were able to grow into plants. They included embryos arrested at the late torpedo and cotyledonary stages, and an arrested torpedo embryo that resulted from precocious hypocotyl expansion. Early torpedo-stage embryos rooted on reculture but did not form shoots. A diversity of other embryo stages were also produced which upon reculture formed only adventitious embryos and roots. Morphologic variants with similar growth potential were noted among embryos at similar stages of development. Plasticity in somatic embryo developmental patterns led to different forms that were mature enough to produce plants.

Endo-β-mannanase activity and seed germination of thermosensitive and thermotolerant lettuce genotypes in response to seed priming

The effects of seed priming on germination and endo-β-mannanase activity at inhibitory and non-inhibitory temperatures in thermosensitive ‘Dark Green Boston’ (DGB) and in thermotolerant ‘Everglades’ (EVE) lettuce were investigated. A single-seed endo-β-mannanase assay was used to follow the enzyme activity during priming. Seeds were primed at 15°C in aerated solutions of polyethylene glycol (PEG) with constant light, then redried. Primed and non-primed seeds germinated 100% at 20°C. At 35°C, non-primed and primed EVE seeds germinated 100%, whereas non-primed seeds of DGB germinated only 4%. During priming, endo-β-mannanase activity increased between 24 and 48 h in EVE and between 24 and 72 h in DGB after the beginning of osmotic imbibition. Endo-β-mannanase activity persisted in primed seeds following seed drying, was detected before radicle protrusion and was present in the micropylar region in front of the radicle tip. Higher enzyme activity was observed in primed seeds and EVE compared with non-primed and DGB seeds. The results suggest that priming may overcome the inhibitory effect of high temperature in thermosensitive lettuce seeds due to increased endo-β-mannanase activity, possibly leading to a weakening of endosperm, thus overcoming thermodormancy.

Effect of Age of Cocomposted MSW And Biosolids on Weed Seed Germination

The influence of municipal solid waste (MSW) and biosolids compost maturity on germination of several weed species seeds was evaluated. Ivyleaf morning glory (Ipomoea hederacea L.), barnyard grass (Echinochloa crus-galli L.), common purslane (Portulaca oleracea L.), and corn (Zea mays L.) were selected as plant indicators to determine the compost maturity stage with maximum germination inhibition. Extracts were prepared from immature (three day-old, four week-old, eight week-old), and mature (one year-old) composts. Extract from eight week-old compost decreased percentage germination, root growth, and germination index (a combination of germination percentage and root growth), and increased mean days to germination of each indicator specie. Extract from eight week-old compost was evaluated for effect on germination percentage of 14 economically important weed species. Extract from eight week-old compost inhibited germination of most weed species, except yellow nutsedge (Cyperus esculentus L.) for which tu...