Documentos de Académico
Documentos de Profesional
Documentos de Cultura
Need of new breeding tools The total population of the world is more than 7 billions & increases with the 1.2% growth rate per year. Food demand increases with more than 3.2% per year. Area of agricultural is decreasing rapidly every year. Conventional plant breeding has various limitations. Feeding the entire population is a major challenge for plant breeders.
So, Whats the way out ? Molecular base tools. - MAS, etc. Tissue culture base tools Shoot tip & meristem culture Root cultures Protoplast cultures Haploid/Doubled haploid culture, etc.
Haploid/Doubled haploid
History
D. Bergner (1921):- Observed first haploid plant (natural).
Successful crops in which Doubled haploid lines were developed: Rape (52), Wheat (36), Rice (11), Melon (9) Pepper (8), Asparagus (7), Tobacco (6), Eggplant (5), Triticale (3), Turnip (1), etc.
Induction of haploidy
Androgenesis
Gynogenesis
Androgenesis
It is most popular & attempted for many species. Use male gametophyte (microspore or immature pollen). Through a series of cell division and differentiation develop haploids. It is most frequent in family Solanaceae and Poaceae. It is of two types:-
A) Direct Androgenesis Normal embryogenesis and form embryo & then plant. Frequent in Solanaceae, Brassicaceae, etc.
B) Indirect Androgenesis Divide repeatedly to form callus & then differentiates into plants. Frequent in barley, wheat, Maize, coffee, etc.
o Methodologies
Anther Culture Microspore/Pollen Culture
Anther Culture: Culture the developing anthers at a precise and critical stage of unopened flower bud. The microspores develop into callus tissue or embryoids. Give rise to haploid plantlets.
Microspore/Pollen Culture Culture the uninucleate Microspore/Pollen. The microspores develop into callus tissue or embryoids. Give rise to haploid plantlets.
Separate Anthers
Callus or Embryo
Incubation
Regeneration
Incubation
Plantlet (n)
o Pathway of development:- Divisions in responding pollen grains may occur in four different pathways:
The pollen grain divide symmetrically Two equal daughter cells undergo further divisions.
The pollen divides unequally. The vegetative cell divides in to callus or embryo.
The pollen divides unequally. The generative cell divides in to callus or embryo.
The pollen grains divide unequally,. The generative and vegetative both cells divides in to callus or embryo.
Gynogenesis
Alternating way to Androgenesis.
First report on the induction of gynogenic haploids was in Barley by San Noeum (1976)
Use female gametophyte (embryo sac-ovary and ovule). But more monotonous than androgenesis. Mostly useful where androgenesis has been unsuccessful. It is mostly used in barley, rice, wheat, etc.
o Methodologies
Ovary Culture Ovule Culture
Ovary Culture:Culture the unpollinated ovaries of mature flower bud. The ovary develop into callus tissue or embryoids. Give rise to haploid plantlets.
Ovule Culture Culture the ovules of mature flower bud. The ovules develop into callus tissue or embryoids. Give rise to haploid plantlets.
Separate Ovary/Ovule
Inoculate on culture medium
Incubation
Callus or Embryo
Incubation
Regeneration
Incubation
Plantlet (n)
Ovary
Ovule
Parthenogenesis
The egg cell of embryo sac typically develops in to embryo without involment of the sperm nucleus. It is called as pseudogamy.
It is induced Invivo by chemical treated pollen (cobalt 60), followed by Invitro culture of embryogenesis.
Chemical treatment prevents pollen fertilization and stimulates the development of haploids from ovules. Useful in those species in which anther culture has not been successful. Frequency of parthenogenetic haploid development is too low.
Culture Medium
Common Media
Murashige and Skoog (MS) medium.
Growth medium
Organic supplement
Vitamins (B1)
Gelling Agent
Agar
Gelrite
Myo-inositol Sucrose Phytagel Agarose, etc.
Complex organics Coconut milk Coconut water Yeast extract Fruit juices Fruit pulps etc.
Micronutrients
Iron(Fe) Manganese (Mn) Zinc (Zn) Boron (B) Cupper(Cu) Molybdenum(Mo) Cobalt (Co) Iodine (I)
Growth Regulators
Auxin Cytokinin
Gibberelin
Classification
Abscisic acid Jasmonates
Ethylene Other
Salicylic acid
Brassinosteroids
GR
Examples
Effects
Indole-acetyl-L- alanine
Auxins Indole-acetyl-L-glycine
2,4Dichlorophenoxyacetic
acid (2,4-D)
bending in response to
gravity or light
GR
Effects Stimulate cell division Stimulate shoot initiation and growth of lateral buds Stimulate dark germination Stimulate leaf expansion
GR
Examples
Diterpenes synthesized via the mevalonic acid
Effects
Stimulate stem elongation by stimulation cell division and elongation Break seeds dormancy Stimulate germination of pollen
Gibberellins
pathway They are more than 110 and named as GA1, GA2, GA3...GA110
GR
Examples
Effects
It is a sesquiterpene Involved in the abscission Abscisic Acid (ABA) of buds, flower and fruits Inhibit cell division and elongation It is a gas produced Regulate ripening of fruits Ethylene from L-methionine Inhibit flowering Regulate cell death programming
GR
Effects
Promote flowering
Salicylic acid Stimulate plant pathogenesis protein production Jasmonate Play an important role in plant defence mechanisms
4) Culture Medium: Medium requirements vary with genotype, age of anther, etc. Growth regulators activated charcoal, iron stimulate the induction of androgenesis and gynogenesis.
5) Effect of temperature (Low/High): Temperature treatment enhance the haploid induction. Temperature treatment maintenance the higher ratio of viable. pollen.
6) Mutagenic Treatment: Exposure to irradiation (gamma rays) promote androgenesis and gynogenesis.
7) Other additives: PCIB, Colchicine, etc. in the culture medium shows improvement in haploid production.
Diploidization of haploids
Diplodized to produce inbred lines. For diploidization microtubule depolymerizing herbicide have been use Colchicine, Trifluralin, Oryzalin, APM, etc. The diploidization carried out in different method. Apply in to the secondary buds. Submerging the roots of plantlets. Direct treatment to embryo or plantlets. Direct treatment to microspore, etc.