Tissue Culture Techniques for Horticultural Crops

Increasing demand for ferns means that more knowledge of propagation techniques be developed to guarantee the continuous supply of these plants from commercial ornamental companies In vitro tissue culture techniques show a certain promise with respect to a future increase in production This review summarises all the research that has been conducted with ornamental ferns using in vitro techniques at different phases in their life cycle, starting from spores, to gametophytes and sporophytes Nutritional, environmental and other factors affecting growth and development in each phase of the life cycle of the fern are discussed, highlighting some protocols that enhance their morphogenic and reproductive capacity

In vitro culture of ornamental ferns

Vegetative reproduction by the gametophyte generation has allowed a number of fern species to persist beyond their normal geographic range. In at least two of these, Trichomanes and Vittaria in the eastern United States, the sporophyte stage of the life cycle has been eliminated completely, perhaps for ten million years or more. Despite this long period of reliance on vegetative reproduction, genetic diversity in the Appalachian Vittaria gametophytes, as measured by starch gel enzyme electrophoresis, remains comparable to that of sexual plants, although it is largely partitioned into monomorphic populations. Widely separated populations are not particularly di- vergent; the taxon as a whole appears equally as cohesive as sexual species, suggesting that factors other than gene flow are responsible for long term maintenance of species integrity. The Appalachian Vittaria gametophytes and similarly derived taxa merit recognition as distinct species. Sexually reproducing pteridophytes alter- nate a diploid, spore-producing plant, the spo- rophyte, with a haploid, gamete-producing plant, the gametophyte. In most taxa, the ga- metophyte generation is short-lived and incon- spicuous relative to the sporophyte plants, which are generally perennial. Life spans of

http://www.jstor.org/stable/pdfplus/2419020.pdf

Species and Evolution in Asexually Reproducing Independent Fern Gametophytes

In vitro culture response was assessed in tomato ( Lycopersicon esculentum Mill. c. v. Omdurman) for optimum callus induction and plantlet regeneration. Callus induction was achieved within seven to ten days directly on the cut surfaces of both hypocotyls and cotyledon explants cultured on Murashige and Skoog (MS) basal medium supplemented with α-naphthalene acetic acid (NAA), 2,4-dichlorophenoxyacetic acid (2,4-D), Thidiazuron (TDZ) and benzyl adenine (BA) alone or in different combinations, but not in hormone free-medium. The highest callusing index (5.3) was obtained on hypocotyls explants cultured on MS medium supplemented with NAA at 0.5 mg/l followed by an index of 5.2 obtained from the same explant by using 0.1 mg/l NAA in combination with BAP at 0.5 mg/l. However, for the cotyledon explants, the highest callusing index (4.7) was obtained on MS medium supplemented with NAA at either 2.0 or 3.0 mg/l. After 8 weeks of culture, organogenesis was observed only on the explants cultured on medium containing different concentrations of TDZ alone or in combination with BAP. The best shoot formation (93%) was obtained for cotyledon explant callus induced on MS medium containing TDZ in combination with BAP both at 0.5 mg/l. The highest number (6) of shoot per explant was obtained when cotyledon explant callus was sub cultured on MS medium supplemented with 3.0 mg/l TDZ. Plain half strength of MS was found to be the best rooting medium, however, addition of IAA at 1.0 mg/l and IBA at 2.0 mg/l were found essential to induce highest number of roots (22.1 ± 0.9) and longer roots (11.0 ± 0.3 cm), respectively. This protocol would be useful to create somaclonal variation and utilize transgenic approaches for varietal improvement of tomato.

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Callus formation and organogenesis of tomato (Lycopersicon esculentum Mill, C.V. Omdurman) induced by thidiazuron

The present study was conducted to investigate the effects of different concentrations and combinations of growth regulators on callus induction and plant regeneration of potato (Solanum tuberosum L.) cultivar Diamant. The tuber segments were used as explants and cultured on Murashige and Skoog (MS) medium supplemented with different concentrations of -naphthalene acetic acid (NAA), 2,4-Dichlorophenoxy acetic acid (2,4-D), benzyl adenine (BA) and thidiazeron (TDZ) alone and 2,4-D in combinations with BA for callus induction. The best degree for callus formation (6.0) was obtained on MS medium supplemented with 2,4-D alone at 3.0 mg/l or 2,4-D in combination with BA both at 2.0 mg/l. MS media supplemented with different levels of BA and TDZ were employed for shoot regeneration. MS medium containing 5.0 mg/l TDZ was the best for days to shoot initiation, the highest percentage of callus with shoot (81%) and highest number of shoot per callus (3.4). Callus derived shoots were rooted most effectively in half-strength MS medium containing 0.5 mg/l IBA. The success of plant tissue culture for in vitro culture of potato was encouraged by acclimatization of the plantlets in the greenhouse conditions. Regenerated plants were morphologically uniform with normal leaf shape and growth pattern.

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Effect of plant growth regulators on callus induction and plant regeneration in tuber segment culture of potato ( Solanum tuberosum L. ) cultivar Diamant

This study is concerned with the determination of the shoot and root inducing effects of kinetin (K) and naphthaleneacetic acid (NAA) on shoots of the fishtail fern (Nephrolepisfalcata forma furcans) in sterile tissue culture. The data shows that K is the major factor involved in maximal shoot production. The NAA is not essential. Specific concentrations of NAA must be present with specific concentrations of K for maximal root production. At the same time, the data also demonstrate that shoots on media containing no NAA but with K concentrations of 5 X 10-7 and 10-6 M produced as many roots as with any other NAA concentration. The data can be used as a guide to rapid commercial propagation of fishtail fern, and demonstrate that media available commercially for Boston fern multiplication will induce maximal shoot production in fishtail fern. MURASHIGE (1974) developed media and techniques for the rapid clonal multiplication of several herbaceous plants using aseptic techniques. Murashige's review is the first publication that refers to the in vitro propagation of any of the members of the genus Nephrolepis. However, specific data collected for the specific plants in that review were unpublished since the research was done in his laboratory. A more detailed study on Nephrolepis exaltata bostoniensis, the popular Boston fern, was done by one of his former graduate students (Harper,

The effects of kinetin and naphthaleneacetic acid on in vitro shoot multiplication and rooting in the fishtail fern.

Asexually-reproducing gametophytes of the four tropical fern genera Hymenophyllum, Trichomanes, Grammitis, and Vittaria occur in the uplands of the southeastern United States (Farrar, 1967). The most common and abundant of these is the Appalachian Vittaria gametophyte, which Farrar (1978) has described in detail. Dr. A. J. Sharp of the University of Tennessee was the first to find and collect the Appalachian Vittaria gametophyte in the area of the Mountain Lake Biological Station in Virginia. Since that time, Dr. W. H. Wagner, Jr. of the University of Michigan collected it in the same area and catalogued it into a pteridophyte flora (Wagner, 1963). Other pteridologists have collected it in the area of the Highlands Biological Station in North Carolina (Wagner et al., 1970; Pittillo et al., 1975) and in Tennessee (Wofford & Evans, 1979). Other collections have located these gametophytes in several eastern states in and around the southern Appalachians (Farrar, 1978; Gastony, 1977). Appalachian Vittaria gametophytes are long-lived and produce extensive colonies in deep, protected crevices of non-calcareous rock. Although antheridia and archegonia are produced on the gametophytes, reduced, juvenile sporophytes have been found only once in the field by Farrar (1978). They were recognizable as Vittaria, but lacked sufficient characters for identification to species. Since most taxonomic characters for determining fern species are based on sporophyte morphology, the origin and affinities of these gametophytes are uncertain. They were first assigned to the genus Vittaria by Wagner and Sharp (1963) based on finding similar morphological characters between the gametophytes of the Appalachian Vittaria and those of Vittaria lineata (L.) J. E. Smith from Florida. The chromosome counts reported by Gastony (1977) have provided further evidence of their relationship. The experimental production of sporophytes might provide additional taxonomic characters for comparison with the Vittaria species. Farrar (1978) reported that Appalachian Vittaria gametophytes were difficult to maintain in axenic culture and ceased to grow on defined media. We report here successful axenic culture of the Appalachian Vittaria gametophyte and the spontaneous formation of callus and sporophytes.

Michael J. Beck

Papers

The effects of kinetin and naphthaleneacetic acid on in vitro shoot multiplication and rooting in the fishtail fern.

Axenic Culture and Induction of Callus and Sporophytes of the Appalachian Vittaria Gametophyte