Michael E. Compton

Use of tissue culture and biotechnology for the genetic improvement of watermelon

Watermelon is an important vegetable crop world-wide with over 81 million metric tons produced annually. Despite these high production figures, million of metric tons of fruit are lost in fields to disease. Genetic improvement through tissue culture and biotechnology offer potential routes of improving fruit harvest by offering higher quality products, like seedless fruit, or by introducing recombinant genes or generating somaclonal variants with improved resistance to biotic or abiotic stresses. The purpose of this review is to highlight how tissue culture and biotechnology have been used for the genetic improvement of watermelon and provide suggestions for future application of these methods to facilitate further genetic improvement.

Statistical considerations for in vitro research : I-Birth of an idea to collecting data

SummaryThis manuscript is the first of two that discuss statistical methods applicable to in vitro research. This paper focuses on topics to consider when planning experiments and provides examples of several experimental designs that are suitable for plant tissue culture and biotechnology projects. Use of factorial treatment designs and designing sequential experiments are also discussed. The second paper deals with data handling. Several key references are provided at the end of each manuscript as sources of additional information for readers’ perusal.

Statistical considerations for in vitro research: II — Data to presentation

SummaryThe paper is the second of two papers about statistical considerations that researchers should make while doing in vitro plant biology research. The first paper focused on aspects from developing a plan to do research through the collection of data. This paper continues with information about editing data, handling outliers, analyzing quantitative and qualitative data, comparing treatment means, preparing graphs and tables, and presenting results.

Micropropagation for recovery of Cucumis hystrix

A micropropagation protocol that allows for the efficient cloning of C. hystrix was developed using 1 mm shoot-tip explants prepared from plants grown in the greenhouse. Establishment of Stage I cultures was greatest (83%) when shoot tips were cultured in modified MS (Murashige and Skoog, 1962) medium containing (per liter) 30 g sucrose, 0.1 g myo-inositol, and 5 g Agargel plus 1.7 μM indole-3-butyric acid (IBA), 0.5 μM kinetin and 0.3 μM gibberillic acid (GA3) (IKG). Benzyladenine (BA, 5 μM) proved best for Stage II shoot proliferation. Over 35 shoots were obtained per vessel (five shoots per vessel) when explants were cultured in medium containing BA. Less than 10 shoots were obtained per vessel when kinetin (0.5–5 μM) and IKG were used. Stage II cultures established from 1 cm shoot tips obtained from Stage I shoots produced more shoots (1.3-fold) than single node explants. Rooting and plantlet acclimatization were best when shoots greater than 1.5 cm were incubated in MS medium containing 0.5 μM naphthalene acetic acid (NAA). Higher NAA concentrations stimulated rooting but inhibited shoot elongation and reduced the ability of plantlets to survive acclimatization to ambient conditions. Plantlet height influenced acclimatization. Over 72% of plantlets survived acclimatization if they were at least 4.6 cm when transferred to soilless growing medium.

Influence of plant preservative mixture (PPM)TM on adventitious organogenesis in melon, petunia, and tobacco

SummaryThe influence of PPMTM on somatic embryogenesis in melon, adventitious shoot organogenesis in petunia, and androgenesis in tobacco was studied by culturing explants in regeneration media supplemented with 0, 2, 5 or 10 ml l−1 PPM for 8–12 wk. The percentage of melon cotyledon explants that produced callus and somatic embryos and the number of embryos per explant were reduced when incubated in embryo initiation and embryo development media containing more than 5 ml l−1 PPM. Less PPM was required to inhibit petunia shoot organogenesis. The number of shoots and number of buds per Petri dish were reduced 3–6.9-fold when leaf explants were incubated in shoot regeneration medium containing more than 2 ml l−1 PPM. In contrast, the addition of up to 10 ml l−1 PPM to tobacco anther culture medium had no effect on androgenesis. Our results suggest that the influence of PPM on plant regeneration depends on the plant species. We recommend that experimenters examine a range of PPM concentrations when using it for the first time on an untested plant species.

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.

Genetic Transformation of Watermelon

Watermelon, Citrullus lanatus (Thunb.) Matsum. and Nakai, is an important vegetable crop globally because of its high vitamin [25% and 20% of the USA recommended daily allowance (RDA) of vitamins C and A, respectively, per 0.280 g fwt.] and nutrient content (8% US-RDA of potassium, 4% US-RDA iron and 2% US-RDA of calcium per 280g fwt.). Watermelon flesh is also high in lycopene, a potent antioxidant that has been shown to reduce human risk to cancer of the prostrate, pancreas and stomach (1, 2).

Use of statistics in plant biotechnology.

Statistics and experimental design are important tools for the plant biotechnologist and should be used when planning and conducting experiments as well as during the analysis and interpretation of results. This chapter provides some basic concepts important to the statistical analysis of data obtained from plant tissue culture or biotechnology experiments, and illustrates the application of common statistical procedures to analyze binomial, count, and continuous data for experiments with different treatment factors as well as identifying trends of dosage treatment factors.