Chemistry: molecular biology and microbiology – Plant cell or cell line – per se ; composition thereof;... – Culture – maintenance – or preservation techniques – per se
Reexamination Certificate
1997-05-22
2001-06-05
Marx, Irene (Department: 1651)
Chemistry: molecular biology and microbiology
Plant cell or cell line, per se ; composition thereof;...
Culture, maintenance, or preservation techniques, per se
C435S430000, C435S430100, C435S431000
Reexamination Certificate
active
06242257
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to a tissue culture process for producing a large number of viable cotton plants in vitro from a specified tissue of cotton plant. The invention provides genotype independent, direct, multiple shoot proliferation and opens up new possibilities for micropropagation, selection of mutants and for producing genetically improved cotton plants by modern methods of agrobiotechnology and genetic engineering. The protocol provides an important step in the success of cotton improvement programmed, utilizing tissue culture technology.
BACKGROUND AND PRIOR ART REFERENCES
Cotton is a globally important crop, grown primarily for fiber. Seeds provide an important source of food for livestock. Cotton has influenced economic development of many nations, throughout the world. Therefore, cotton improvement programmes by modern methods of agrobiotechnology are of interest worldwide. This has increased the importance of developing tissue culture methods to facilitate the application of modern techniques of genetic engineering of cotton plant.
Several reports on tissue culture of cotton have been published. These are related to direct shoot differentiation and somatic embryogenesis through suspension and callus cultures. These are listed below by way of references.
Organogenesis and regeneration, leading to micropropagation by tissue culture methods have been successfully demonstrated in several plant species e.g. Phaseolus sp. (Rubluo A & Kartha K K 1985). In vitro culture of shoot apical meristems of various Phaseolus species and cultivars, J. of Plant Physiol 119: 425-433
. Glycine max
(Shetty K., Asano Y and Oosawa K 1992 Stimulation of in vitro shoot organogenesis in
Glycine max
Merrill by allantoin and amides. Plant Sci. 81:245-252).
Cajanus cajan
(Shiv Prakash N, Pental D & Bhalla-Sarin N 1994 Regeneration of Pidgeonpea (
Cajanus cajan
) from cotyledonary node via multiple shoot formation. Plant Cell Rep. 13:623-622), Carnation (Claire Annex A. Yancheva S and Dons H 1995. Cells within the nodal region of carnation shoots exhibit a high potential for adventitious shoot formation. (Plant Cell Tiss. Org. Cult. 40:151-157) etc.
Plant regeneration by tissue culture techniques is well established. A wide variety of plant species has been successfully regenerated in vitro via organogenesis or somatic embryogenesis. Organogenesis leads to organ formation i.e., shoot (or root), which can be isolated to induce development of roots (or shoots) to produce full plant while somatic embryogenesis leads to the development of somatic embryos (embryos developed without fertilization) which have both shoot and root initials and are capable of developing into whole plant. Although the ability of individual parts of plants and cells to regenerate into complete plants (called totipotency) is a well known phenomenon, each plant or plant part requires specialized studies to invent the conditions that allow such regeneration. Some of the broadly applicable factors controlling growth and differentiation of such cultures have been determined. The establishment of interactions among different groups of phytohormones and growth regulators alone or in combinations are responsible for certain interrelations existing among cells, tissues and organs. There seems to be consensus that the success in inducing differentiation depends upon the type of explant, physiological condition of the explant and physical and chemical milieu of the explant during culture. Due to this, the science of tissue culture has been directed to optimize the physiological conditions of source plant, the type of explant, the culture conditions and the phytohormones used to initiate tissue culture. This substantiates the fact that development of a new process for proliferation of plants by tissue culture is not obvious.
One major aspect that has to be investigated on case to case basis is the type of plant growth regulators and the amount of plant growth regulators that induce regeneration. Besides chemical composition of the medium, temperature of growth and other culture conditions play important role in the induction of organogenesis and somatic embryogenesis and maturation of shoots and roots and the formation of healthy fertile plants. The response to medium, hormones and growth conditions differs from plant species to species and variety to variety. Thus, inventing conditions for efficient regeneration of plants, organogenesis and somatic embryogenesis requires developing specialized knowledge about a given plant.
Another major area where innovativeness is required in tissue culture is identifying the plant part that efficiently responds to the culture conditions and leads to prolific regeneration. Not all plant parts of a given species are amenable to efficient regeneration. It is a complex combination of the plant part identified for totipotency (called explant), the physiological state of the explant and the growth conditions, especially the growth regulators that determine success of a plant in tissue culture. Different explants from a given plant usually show very different response to growth conditions for proliferation. No general principles can be applied to achieve regeneration. In each case, identification of the explant and identification of the culture conditions are innovative steps in the development of a tissue culture method for regeneration of a plant part into a number of plants or somatic embryos.
As of now, a detailed publication on the formation of multiple shoots from any tissue explant of cultivated varieties of cotton is not available. This invention describes for the first time a detailed protocol for organogenesis from a small part (called an explant) of cotton seedling to give multiple shoots of cotton plant through tissue culture. The method is very useful in agricultural biotechnology for micropropagation and genetic transformation because it shows wide applicability to all the cultivars tested by the inventors and the shoots can be efficiently raised to maturity. The method has a great potential in cotton improvement programmes by modern methods of agrobiotechnology.
Several reports deal with tissue culture conditions (Davidonis G H and Hamilton R H, 1983 Plant regeneration from callus tissue of
Gossypium hirsutum
L. Plant Sci. Lett. 32: 89-93; Shoemaker R C, Couche L J & Galbraith D W 1986). Characterization of somatic embryogenesis and plant regeneration in cotton (
Gossipium hirsutum
L) Plant Cell Rep. 3:178-181; Trolinder N L and Goodin J R 1987; Somatic embryogenesis and plant regeneration in cotton
Gossipium hirsutum
L. Plant Cell Rep. 6:231-234 Finer J. 1988). Plant regeneration from somatic embryogenesis suspension cultures of cotton
Gossipium hirsutum
. (Plant Cell Rep. 7:399-402) that give rise to somatic embryos (structures that give rise to normal plants without going through fertilization) have been reported. Initiation and maturation of somatic embryos take several months. The method is highly dependent on genotypes (Trolinder N L and Xhixian C., 1989 Genotype specificity of the somatic embryogenesis response in cotton. Plant Cell Rep. 8:133-136) and is, therefore, applicable to a restricted group of varieties. Initiation and maturation of somatic embryos takes several months, and the plants regenerated via somatic embryogenesis were of ten reported to be cytologically and morphologically abnormal (Stelly D M, Altman D W, Kohel Rz, Rangan T S & Commeskey E 1989. Cytogenetic abnormalities of cotton somaclones from callus cultures. (Genome 32:762-770). Plants developed via somatic embryogenesis were also of ten reported to be sterile (Trolinder N L & Goodin J R 1987, Somatic embryogenesis and plant regeneration in cotton
Gossipium hirsutum
L. Plant Cell Rep. 6:231-234).
These regeneration processes have been successfully used in Agrobacterium mediated gene transfer in cotton (Umbeck P, Johnson G, Barton K. Swain W 1987 “Genetically transformed cotton
Gossipium hirsutum
L. Plant Bio./ Technology 5:263-266; Firoozabady E., DeBoer L D, Merlo
Gupta Shiv Kumar
Srivastava Alok Kumar
Tuli Rakesh
Chung Dutch D.
Council of Scientific & Industrial Research
Fish & Neave
Marx Irene
Pierri Margaret A.
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