V G SHOBHANA

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BIOFORTIFICATION IN MAIZE

• V G SHOBHANA
• Dr. N SENTHIL
• KALPANA K.
• Dr. P NAGARAJAN
• Dr. M RAVEENDRAN
• Dr. P BALASUBRAMANIAN

• CENTRE FOR PLANT MOLECULAR BIOLOGY
• TAMIL NADU AGRICULTURAL UNIVERSITY
• COIMBATORE 641 003

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BIOFORTIFICATION
Fortification x Biofortification

• Methods
• Selective Breeding
• Genetic modification
• The Big Difference!!
• Developing world
• Vitamin A, Zinc, Iodine and Iron
• Developed world
• Selenium, prostrate cancer

The Orange Ribbon Symbol of Malnutrition
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Importance

• Two billion people - currently micronutrient
  malnourished - increased morbidity and mortality
  rates, lower worker productivity and high
  healthcare costs.
• Nutritional deficiencies (iron, zinc, vitamin A)
  - almost two-thirds of the childhood death
  worldwide.
• Major food crops can be enriched
  (biofortified) with micronutrients using plant
  breeding and transgenic strategies.
• Micronutrient enrichment traits exist within
  their genomes.
• Micronutrient element enrichment of seeds can
  increase crop yields when sowed to
  micronutrient-poor soils, assuring their adoption
  by farmers.

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The Golden Rice Story
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Percentage of population affected by
under-nutrition by country, according to United
Nations statistics
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Phytic acid (Phytin or Phytate)
BIOSYNTHETIC PATHWAY

• Myo-inositol-1,2,3,4,5,6-hexakisphosphate or Ins
  P6.
• Is the most abundant myo-inositol phosphate in
  plant cells, but its biosynthesis is poorly
  understood.
• Also uncertain is the role of myo-inositol as a
  precursor of phytic acid biosynthesis.
• MW 660.03Formula C6H18O24P6

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• PHYTIC ACID
• Myo-inositol 1,2,3,4,5,6-hexakisphosphate, is
  abundant component of plant seeds.
• Deposited in protein bodies as a mixed salt of
  mineral cations such as K, Mg2, Ca2, Zn2,
  and Fe 3 (50 to 80 of the phosphorus in
  seeds).
• Phytic acid serves as a major storage form for
  myo-inositol, phosphorus, and mineral cations for
  use during seedling growth.
• Other known role of phytic acid - control of
  inorganic phosphate (Pi) levels in both
  developing seeds and seedlings.
• In maize kernels, nearly 90 is accumulated in
  embryo and 10 in aleurone layers (also in rice
  and barley).
• Maize endosperm contains only trace amount of
  phytic acid.

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Importance

• Monogastric animals digest phytic acid poorly.
• Undigested phytic acid is eliminated and is a
  leading phosphorus pollution source.
• Low-phytic acid grain and legume in feed -
  reduces phosphorus pollution to environment and
  reduce amount of phosphorus supplementation
  required in animal feeds (Ertl et al., 1998).
• Such grain would also offer more available Fe and
  Zn for human nutrition (Mendoza et al., 1998).

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Variability of phytate P in crop plants
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Biosynthetic pathways of phytate in plants

• Two types of pathway
• Lipid -dependent (hydrolysis of PI(4,5)P2 by
  phospholipase)
• Lipid -independent (sequential
  phosphorylation of I(3)P or inositol)

Paulik et al.,(2005)
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Analysis of biochemical characters

• Phytic acid Wheeler and Ferrel, 1971
• 430 genotypes were screened for their phytate
  content
• Low and high maize inbreds were identified
• Crossing of low inbred with high inbreds evolved
  in 50 hybrids
• Iron and Zinc major minerals screened by
  Atomic Absorption Spectrophotometer

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The following strategies were adopted to reduce
the phytate

• Plants can be transformed for increased phytase
  production in the seeds.
• The transgenic approach will, in the long run,
  prove to be most versatile and cost-effective.
• Mutation breeding for impaired phytic acid
  biosynthesis has proved to be useful in maize,
  barley and rice ( Raboy, 2000).
• Available low phytate mutant lines can be crossed
  with locally adopted cultivars and will result in
  low phytate maize with desired agronomic
  backgrounds.

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Genetics

• Maize has 10 chromosomes (n10).
• The combined length of the chromosomes is 1500
  cM.
• "Chromosomal knobs". They are highly repetitive
  heterochromatic domains that stain darkly.
• Barbara McClintock used these knob markers to
  prove her transposon theory of "jumping genes".

Composition Figures in grams (g) or milligrams
(mg) per 100g of food.
Minerals Calcium 9mg Phosphorus 290mg Iron 2.5m
g
Seed (Fresh weight) 361 Calories per 100g
Water 10.6 Protein 9.4g Fat 4.3g Carbohydrate
 74.4g Fiber 1.8g Ash 1.3g
Vitamins Vit A 140mg Thiamine
(B1) 0.43mg Riboflavin (B2) 0.1mg Niacin 1.9mg
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Mutation work - Dr. V Raboy, USDA

• Pollen treated M2 progenies - developed by Dr.
  Raboy yielded two maize mutants.
• lpa 1 and lpa 2 with 60 reduction in the seed
  phytate levels were produced.
• These mutants were widely used in most of the
  breeding programmes in US.
• lpa 1 1.1 (mg/g) phytate P in 4.7 (mg/g) total
  P
• lpa 1 2.6 (mg/g) phytate P in 4.6 (mg/g) total
  P
• Indian corns have 2.0 2.5 (mg/g) phytate P in
  4.0 - 4.5 (mg/g) of total P.

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INBREDS SELECTED FOR MUTATION BASED ON PHYTIC
ACID CONTENT
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INBREDS SELECTED FOR MUTATION BASED ON THE PHYTIC
ACID CONTENT
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Methodology

• Low phytic acid donors with lpa1 and lpa2 genes
  will be used from Victor Raboy, USDA and will be
  used to develop low phytate maize.
• Local inbred lines will be used as recurrent
  parents.
• Identification of closely linked DNA markers with
  phytate in maize using already available linked
  markers like umc157 with lpa1 and umc167 with
  lpa2.
• Develop backcross population and marker assisted
  backcross selection for low phytate maize lines.

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Expected output

• Identification of low phytate genotypes of maize
  which could be potential donors in breeding for
  micronutrients.
• Molecular markers linked to low phytate will
  assist in identifying target genes involved in
  adsorption, transport and unloading of
  micronutrients in the grain.
• Low phytate versions of high yielding maize
  hybrids in cultivation in India with increased
  iron and zinc bioavailability and reduced
  phosphorus pollution in the environment.