tissue culture Plant tissue culture is a technique of growing plant cells tissues organs seeds or other plant parts in a sterile environment on a nutrient medium WHY The production of ID: 932568
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Slide1
Plant Tissue Culture
Slide2What is plant tissue culture
?
Plant tissue culture is a technique of growing plant cells, tissues, organs, seeds or other plant parts in a sterile environment on a nutrient medium
Slide3WHY?
The production of
clones of plants that produce particularly good flowers, fruits, or have other desirable traits.To quickly produce mature plants.
The production of multiples of plants in the absence of seeds or necessary pollinators to produce seeds.The regeneration of whole plants from plant cells that have been genetically modified.
Slide4WHY?
The
production of plants in sterile containers reduces disease transmissionAllows production of plants from seeds that otherwise have very low chances of germinating and growing, i.e.: orchids and
Nepenthes.To clean particular plants of viral and other infections and to quickly multiply these plants as 'cleaned stock' for horticulture and agriculture.
Slide5Terminology
Explant
Living tissue transferred
from a plant to an artificial
medium for culture.
It can be any portion of
the shoot, leaves, roots,
flower or cells from a plant.
Slide6How?
Adult plant cells are
totipotent, meaning they have the ability to give rise to a fully differentiated plant. Because of this, it is possible to collect cells from a mature plant and use those cells to produce clones of that plant.
Slide7Plant tissue Culture Basics
Modern plant tissue culture is performed under
aseptic conditionsLiving plant materials from the environment are naturally contaminated on their surfaces (and sometimes interiors) with microorganisms, so surface sterilization of starting material (
explants) in chemical solutions (usually alcohol and sodium or calcium hypochlorite is required).
Slide8Plant tissue Culture Basics
Explants
are then usually placed on the surface of a solid culture medium, but are sometimes placed directly into a liquid medium, when cell suspension cultures are desired.
Culture media are generally composed of inorganic salts plus a few organic nutrients, vitamins and plant hormones.
Slide9Plant Tissue Culture Media
Slide10Salt Mixtures
Organic Substances
Natural Complexes
Inert Supportive Materials
Growth Regulators
Slide11Macro-nutrient salts
What the ?
NH4NO3 Ammonium nitrate
KNO3 Potassium nitrate
CaCl2 -2 H2O Calcium chloride (Anhydrous)
MgSO4 -7 H2O Magnesium sulfide (Epsom Salts)
KH2PO4 Potassium hypophosphate
FeNaEDTA Fe/Na ethylene-diamine-tetra
acetate
H3BO3 Boric Acid
MnSO4 - 4 H2O Manganese sulfate
ZnSO4 - 7 H2O Zinc sulfate
KI Potassium iodide
Na2MoO4 - 2 H2O Sodium molybdate
CuSO4 - 5 H2O Cupric sulfate
CoCl2 - H2O Cobaltous sulfide
Slide12Macronutrient salts
Nitrogen
– Influences plant growth rate, essential in plant nucleic acids (DNA), proteins, chlorophyll, amino acids, and hormones.
Phosphorus – Abundant in meristematic and fast growing tissue, essential in photosynthesis, respiration.
Potassium
–
Necessary for cell division, meristematic tissue, helps in the pathways for carbohydrate, protein and chlorophyll synthesis.
Slide13Macronutrient salts
Calcium
-
Involved in formation of cell walls and root and leaf development. Participates in translocation of sugars, amino acids, and ties up oxalic acid (toxin).
Iron
-
Involved in respiration , chlorophyll synthesis and photosynthesis.
FeNaEDTA
=
sodium salt of
EDTA sequesters iron, making it available to plants
.
Magnesium -
Involved in photosynthetic and respiration systems. Active in uptake of phosphate and translocation of phosphate and starches.
Slide14Micronutrient salts
Sulfur
-
Involved in formation of nodules and chlorophyll synthesis, structural component of amino acids and enzymes.
Manganese
-
Involved in regulation of enzymes and growth hormones. Assists in photosynthesis and respiration.
Slide15Micronutrient salts
Molybdenum
-
Involved in enzymatic reduction of nitrates to ammonia. Assists in conversion of inorganic phosphate to organic form.
Zinc -
Involved in production of growth hormones and chlorophyll. Active in respiration and carbohydrate synthesis.
Boron
- Involved in production of growth hormones and chlorophyll. Active in respiration and carbohydrate synthesis.
Copper -
Involved in photosynthetic and respiration systems. Assists chlorophyll synthesis and used as reaction catalyst.
Organic Compounds
Carbon Sources – Sucrose, sometimes Glucose or Fructose (Plants Need Carbon)
Vitamins –
Adenine – part of RNA and DNAInositol – part of the B complex, in phosphate form is part of cell membranes, organelles and is not essential to growth but beneficial
Thiamine –
essential as a coenzyme in the citric acid cycle.
Slide17Still other organics
Organic Acids
Citric acid (150 mg/l) typically used with ascorbic acid (100 mg/l) as an antioxidant.Can also use some of Kreb Cycle acidsPhenolic compoundsPhloroglucinol - Stimulates rooting of shoot sections
Slide18Natural Complexes
Coconut endosperm
Fish emulsionProtein hydrolysatesTomato juiceYeast extracts
Malt extractPotato agar
Slide19Growth regulators
auxin - Roots
cytokinin - Shootsgibberellin – Cell Enlargementabscisic acid – Plant stress hormoneethylene – BAD!
Slide20Auxins
Callus formation, rooting of cuttings, and the induction of adventive embryogenesis
IAA
IBA
NAA
2,4-D
2,4,5-T
Picloram
Slide21Cytokinins
-Enhances adventitious shoot formation
BA 2iP KinetinZeatin
Slide22Gibberellin
Not generally used in tissue culture
Tends to suppress root formation and adventitious embryo formation
Slide23Abscisic Acid
Primarily a growth inhibitor but enables more normal development of embryos, both zygotic and adventitious
Slide24Ethylene
Question is not how much to add but how to get rid of it
in-vitroNatural substance produced by tissue cultures at fairly high levels especially when cells are under stress
Enhances senescenseSupresses embryogenesis and development in general
Slide25Hormone Combinations
Callus development
Adventitious embryogenesisRooting of shoot cuttingsAdventitious shoot and root formation
Slide26Slide27Applications[
edit]
Plant tissue culture is used widely in the plant sciences, forestry, and in horticulture. Applications include:The commercial production of plants used as potting, landscape, and florist subjects, which uses meristem and shoot culture to produce large numbers of identical individuals
.1.To conserve rare or endangered plant species.[6
]
2.A
plant breeder may use tissue culture to screen cells rather than plants for advantageous characters, e.g. herbicide resistance/tolerance
.
3.Large-scale
growth of plant cells in liquid culture in
bioreactors
for production of valuable compounds, like
plant-derived secondary metabolites
and
recombinant proteins
used as
biopharmaceuticals
.
[7
]
4.To
cross distantly related species by
protoplast fusion
and regeneration of the novel
hybrid
.
]
Slide285.To
rapidly study the molecular basis for physiological, biochemical, and reproductive mechanisms in plants, for example in vitro selection for stress tolerant plants.
[8]6.To cross-pollinate distantly related species and then tissue culture the resulting embryo which would otherwise normally die (Embryo Rescue
).7.For chromosome doubling and induction of polyploidy,
[9]
for example doubled haploids,
tetraploids
, and other forms of polyploids. This is usually achieved by application of antimitotic agents such as colchicine
or
oryzalin
.
8.As
a tissue for transformation, followed by either short-term testing of genetic constructs or regeneration of
transgenic
plants
.
9.Certain
techniques such as meristem tip culture can be used to produce clean plant material from
virused
stock, such as sugarcane
[10]
, potatoes and many species of soft fruit.
Production of identical sterile hybrid species can be obtained
.
10.Large
scale production of artificial seeds through
somatic
embryogenesis
[11