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 UNIVERSITYCOIMBATORE – 641 003

BIOFORTIFICATION IN MAIZE

Methods: Selective Breeding Genetic modification

The Big Difference!! Developing world –

Vitamin A, Zinc, Iodine and Iron

Developed world – Selenium, prostrate cancer

The Orange RibbonSymbol of Malnutrition

BIOFORTIFICATION

Fortification x Biofortification

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.

Importance

The Golden Rice Story

Percentage of population affected by under-nutrition by country, according to United Nations statistics

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

BIOSYNTHETIC PATHWAY

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.

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).

Importance

Variability of phytate P in crop plants

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)

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

o Plants can be transformed for increased phytase production in the seeds.o The transgenic approach will, in the long run, prove to be most

versatile and cost-effective.o Mutation breeding for impaired phytic acid biosynthesis

has proved to be useful in maize, barley and rice ( Raboy, 2000).o Available low phytate mutant lines can be crossed with locally

adopted cultivars and will result in low phytate maize with desired agronomic backgrounds.

The following strategies were adopted to reduce the phytate

Maize has 10 chromosomes (n=10). 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".

Seed (Fresh weight) 361 Calories per 100g Water: 10.6% Protein: 9.4gFat: 4.3gCarbohydrate: 74.4gFiber: 1.8gAsh: 1.3g

CompositionFigures in grams (g) or milligrams (mg) per 100g of food.

Vitamins Vit A: 140mgThiamine (B1): 0.43mgRiboflavin (B2): 0.1mgNiacin: 1.9mg

MineralsCalcium: 9mgPhosphorus: 290mgIron: 2.5mg

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.

INBREDS SELECTED FOR MUTATION BASED ON PHYTIC ACID CONTENT

INBREDS SELECTED FOR MUTATION BASED ON THE PHYTIC ACID CONTENT

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.

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.