Computer vision analysis of somatic embryos of sweet potato [Ipomoea batatas (L.) Lam.] for assessing their ability to convert to plants.

Apical and axial shoot tips of sweet potato were cultured to produce somatic embryos that mature and develop into plants in basal nutrient medium. However, the lack of high regeneration efficiency is an impediment to the use of somatic embryos to produce synthetic seeds. Conversion experiments with mature embryos over a 20-day period revealed that 80-90% of the embryos formed roots but only 40-50% formed shoots. Using computer vision and canonical or Fisher discriminant function (CDA) analysis along with conversion results, it was possible to correctly classify competent embryos 40-50% of the time based on size features, 50-60% of the time based on shape features, and 55-60% of the time based on color features. Non-competent embryos were correctly classified 65-75%, 55-60%, and 70-75% of the time based on size, shape, and color, respectively. These results can be used effectively to identify and select competent embryos for improved regeneration efficiency.

A comparison of shoot-forming and non-shoot-forming somatic embryos of sweet potato [Ipomoea batatas (L.) Lam.] using computer vision and histological analyses.

Diagnostic structural features for competence to form shoots were tested among sweet potato embryos by combining morphological image capture (using a computer vision system) with anatomical analyses (using light microscopy). Five major morphological variants (`perfect', `near perfect', `limited/no meristematic activity', `disrupted internal anatomy', and `proliferating') were identified among torpedo- and cotyledonary-stage embryos. Among these, only the first two were found to be competent for conversion into plantlets. Lack of organized shoot development in somatic embryos of sweet potato was associated with the following abnormalities: lack of an organized apical meristem, sparcity of dividing cells in the apical region, flattened apical meristem, and multiple meristemoids and/or diffuse meristematic activity throughout the embryo. Diagnostic separation of most shoot-forming and non-shoot-forming torpedo and cotyledonary embryo variants was achieved.

A system and methodology for measuring volatile organic compounds produced by hydroponic lettuce in a controlled environment.

A system and methodology were developed for the nondestructive qualitative and quantitative analysis of volatile emissions from hydroponically grown 'Waldmann's Green' leaf lettuce (Lactuca sativa L.). Photosynthetic photon flux (PPF), photoperiod, and temperature were automatically controlled and monitored in a growth chamber modified for the collection of plant volatiles. The lipoxygenase pathway products (Z)-3-hexenal, (Z)-3-hexenol, and (Z)-3-hexenyl acetate were emitted by lettuce plants after the transition from the light period to the dark period. The volatile collection system developed in this study enabled measurements of volatiles emitted by intact plants, from planting to harvest, under controlled environmental conditions.

Photosynthetic photon flux, photoperiod, and temperature effects on emissions of (Z)-3-hexenal, (Z)-3-hexenol, and (Z)-3-hexenyl acetate from lettuce.

To investigate the effects of environment on plant volatile emissions, 'Waldmann's Green' leaf lettuce was cultivated under different levels of photosynthetic photon flux (PPF), photoperiod, and temperature. A modified growth chamber was used to sample plant volatile emissions nondestructively, over time, and under controlled conditions. Total volatile emission rates were significantly higher from lettuce cultivated under PPF of 360 or 200 micromoles m-2 s-1 compared to 105 micromoles m-2 s-1, and significantly higher under a 16-h photoperiod than an 8-h photoperiod. No differences were detected among emission rates from different temperature treatments. In controlled environments, emissions could be regulated by adjusting environmental conditions accordingly.

Noninvasive evaluation of somatic embryogenesis.

Callus Suspension Cultures of Ipomea batates Poir. cv. White Star were grown in an airlift bioreactor. A machine vision system was used to monitor nondestructively callus growth during a 10 day culture period. Growth data obtained with this system included the overall reactor population and population estimates for the 200-1200-microm fractions at 200-microm intervals. A model of callus growth was developed to explain the mechanics of callus enlargement. The model was based on the assumptions that (1) the calli could not subdivide or shrink, (2) there was a fixed percentage of the initial population for each fraction that was nonviable, and (3) growth rates did not vary with time during the culture period. It was determined that the growth rates and nonviable ratios decreased as fraction size increased.

Optimizing sweet potato [Ipomoea batatas (L.) Lam.] root and plantlet formation by selection of proper embryo developmental stage and size, and gel type for fluidized sowing.

Potassium starch polyacrylamide, potassium acrylate, a copolymer of potassium acrylate and acrylamide, and hydroxyethylcellulose carrier gels were tested to find a fluid drilling material suited for synthetic seeding of sweet potato (Ipomoea batatas (L.) Lam.) somatic embryos. Somatic embryo developmental stage and size, and maturation (incubation) time were also evaluated to improve plantlet formation. All embryos suspended in the fluidized hydroxyethylcellulose gel were viable after six days and 7% developed into plantlets after two weeks. Up to 97% of the somatic embryos suspended in acrylate and/or acrylamide gels died within six days. Root development was at least 10% and plantlet development at least 30% greater when embryos were subcultured on basal medium for 16 instead of 25 days prior to placement and suspension in hydroxyethylcellulose gel. Up to 25% more plantlets were obtained from embryos at the elongated torpedo stage than those at the cotyledonary or torpedo stages of development. When suspended in hydroxyethylcellulose gel embryo length had no effect on the percentage of plantlets obtained.

Dependence of Seed Vigor during Germination on Carbohydrate Source in Endosperm Mutants of Maize.

Differences in seed vigor of four genotypes of maize (Zea mays L.), brittle-1 (bt1), shrunken-2 (sh2), sugary (su), and normal, in an isogenic background, were investigated. Excised whole embryos and axes were grown on Murashige and Skoog (MS) media containing various carbohydrate sources. Of the four genotypes examined, sh2 seeds had the lowest vigor, especially under germination stress conditions. Embryo dry weights of sh2 were less than su and normal but equal to bt1 and made up nearly 25% of the whole seed weight. The sh2 seeds and whole embryos had low starch levels compared with the other three genotypes. Sugar levels were comparable in the three endosperm mutants, which were two times higher than normal. Optimum growth of excised embryos of all genotypes was obtained on MS medium containing 5% sucrose. However, this concentration did not totally overcome poor germination and growth of sh2 embryos and axes. Axes of su and normal had greater growth rates than sh2 and bt1 on sucrose-free medium, although the difference between genotypes decreased when whole embryos were used. When ground endosperm was employed as the carbohydrate source, sh2 embryos germinated and grew poorly, particularly on normal endosperm. With a commercial corn starch as the carbohydrate source, sh2 germlings were shorter in length and displayed a greater loss in dry weight than the other genotypes. The poor growth of sh2 embryos on ground endosperm and starch media may indicate a dysfunction of the scutellum or axis in relation to carbohydrate metabolism and utilization.

Mechanism of seed priming in circumventing thermodormancy in lettuce.

Lettuce (Lactuca sativa L. cv Minetto) seeds were primed in aerated solutions of 1% K(3)PO(4) or water at 15 degrees C in the dark for various periods of time to determine the manner by which seed priming bypasses thermodormancy. Seeds which were not primed did not germinate at 35 degrees C, whereas those which were primed for 20 h in 1% K(3)PO(4) or distilled H(2)O had up to 86% germination. The rate of water uptake and respiration during priming were similar regardless of soak solution. Cell elongation occurred in both water and 1% K(3)PO(4), 4 to 6 h prior to cell division. Both processes commenced sooner in water than K(3)PO(4). Radicle protrusion (germination) occurred in the priming solution at 21 h in water and 27 h in 1% K(3)PO(4).Respiration, radicle protrusion and cell division consistently occurred sooner in primed (redried) seeds compared to nonprimed seeds when they were imbibed at 25 degrees C. Cell division and elongation commenced after 10 h imbibition in primed (redried) seeds imbibed at 35 degrees C. Neither process occurred in nonprimed seeds. Respiratory rates were higher in both primed and nonprimed seeds imbibed at 35 degrees C compared to those imbibed at 25 degrees C, although radicle protrusion did not occur in nonprimed seeds which were imbibed at 35 degrees C. It is apparent that cell elongation and division are inhibited during high temperature imbibition in nonprimed lettuce seeds. Seed priming appears to lead to the irreversible initiation of cell elongation, thus overcoming thermodormancy.

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.

Mechanical Resistance of the Seed Coat and Endosperm during Germination of Capsicum annuum at Low Temperature.

Decoated pepper (Capsicum annuum L. cv Early Calwonder) seeds germinated earlier at 25 degrees C, but not at 15 degrees C, compared to coated seeds. The seed coat did not appear to impose a mechanical restriction on pepper seed germination. Scarification of the endosperm material directly in front of the radicle reduced the time to germination at both 15 degrees C and 25 degrees C.The amount of mechanical resistance imposed by the endosperm on radicle emergence before germination was measured using the Instron Universal Testing Machine. Endosperm strength decreased as imbibition time increased. The puncture force decreased faster when seeds were imbibed at 25 degrees C than at 15 degrees C. The reduction in puncture force corresponded with the ability of pepper seeds to germinate. Most radicle emergence occurred at 15 degrees C and 25 degrees C after the puncture force was reduced to between 0.3 and 0.4 newtons.Application of gibberellic acid(4+7) (100 microliters per liter) resulted in earlier germination at 15 degrees C and 25 degrees C and decreased endosperm strength sooner than in untreated seeds. Similarly, high O(2) concentrations had similar effects on germination earliness and endosperm strength decline as did gibberellic acid(4+7), but only at 25 degrees C. At 15 degrees C, high O(2) concentrations slowed germination and endosperm strength decline.

Parthenocarpic Fruit Development from Grafted Ovaries of Cucuminis sativus L.

Reciprocal cleft and pistillate floral bud grafts were made between parthenocarpic ;Fertilla' and nonparthenocarpic ;MSU 713-5' cucumber (Cucumis sativus L.) lines to localize the site for stimulation of parthenocarpic fruit set. No fruit set on ;MSU 713-5' controls, scion grafted to ;Fertilla,' or rootstock with ;Fertilla' as the scion. ;Fertilla' controls, rootstock, and scions all produced parthenocarpic fruit when grafted to ;MSU 713-5.' When pistillate floral buds of ;MSU 713-5' were grafted to ;Fertilla,' no fruit were produced. However, individual immature pistillate buds of ;Fertilla' developed into mature fruits when grafted onto ;MSU 713-5.' Hence, the immature ovary is the site of stimulation for parthenocarpic fruit set in cucumber.

Morphactin-induced parthenocarpy in the cucumber.

Parthenocarpy was induced in cucumber Cucumis sativus L. by foliar application of morphactin. Morphactin (chlorfluorenol) was most effective in inducing parthenocarpy when applied in the flowering stage, and the response was enhanced by prior treatment with Ethrel (2-chloroethylphosphonic acid). Morphactin stimulated pistillate flower development in a monoecious cultivar but did not modify sex expression of a gynoecious line.