Rice is one of the major cereal crops and the cheapest source of food
energy for 50% of the total world population, predominantly in the developing
countries, but it is deficient in many essential micronutrients (including
iron, zinc, and vitamin A). A diet based on milled rice leads to malnutrition,
with deficiencies being most severe in iron, lysine, iodine, vitamin A,
and zinc (FAO, 1993).
Iron deficiency is the most widespread nutritional disorder in the world.
Its effects on human health are severe, and it is estimated that more
than 3.5 billion people in the developing world are anemic (Ahman et
al. 2000), mostly children and women.
Attempts have been made by different national, international and non-government
organizations to alleviate the severity of iron-deficiency anemia by directly
increasing iron intake through dietary supplementation, fortification,
and food diversification. For various reasons, none of the intervention
strategies has been very successful in reducing the prevalence of irondeficiency
anemia in developing countries. Iron deficiency is therefore a major public
Ferritin is an iron-storage protein found in plants, animals, and bacteria,
which have ferroxidase activity. Increasing the iron content in rice by
introducing the ferritin gene by genetic engineering has been reported
earlier in rice (Goto et al. 1999; Lucca et al.. 2001; Vasconcelos
et al. 2003).
Immature embryos (10-12 days after pollination) and calli derived from
mature seeds were used as transformation material. The plasmid pGPTV-bar/Fer,
which contains the ferritin gene, was introduced into widely cultivated
(in Bangladesh) popular rice variety BRRI Dhan 29, driven by the endosperm-specific
glutelin promoter via the particle bombardment method as described previously
(Vasconcelos et al. 2003). Iron content in the seeds was determined
by the ICP (Inductively Coupled Argon Plasma Spectrometer).
A total of 368 putative primary transformed plants (T0) were
regenerated after bombardment and calli selection. PCR analysis was used
to screen the primary transformants, and eight ferritin-positive
transgenic plants (T0) were obtained. The independent transformation
event and integration pattern of the ferritin gene were confirmed
by Southern blot analysis (Fig. 1). In the subsequent segregation (T1),
the ferritin gene (0.8 kb) was inherited in a typical Mendelian
segregation ratio (3:1). Apart from the expected size of 0.8 kb for the
gene, other larger (>0.8 kb) bands were also observed. However, the
phenotypic expression of the putative transgenic plants appeared to be
healthy in the transgenic greenhouse vis-a-vis the control plants. More
than 95% of the transgenic plants showed normal flowering and produced
T1 seeds obtained from T0 plants were used to analyze
the iron levels of transformed and non-transformed control plants. The
iron content in the polished control seeds was 3.3 mg/kg, whereas it ranged
from 4.5 to 8.9 mg/kg in the transgenic seeds (Fig. 2). Transgenic seeds
of some lines showed about a two fold higher iron content than the control
seeds after polishing.
This is the first report on transferring the soybean ferritin gene
with an endosperm-specific promoter to an elite indica rice cultivar,
BRRI Dhan 29. This finding suggests that the rice lines with enhanced
iron content developed by genetic engineering may help overcome the iron-deficiency
nutritional problem of the population that consumes rice as a staple food.
The work at IRRI has been generously supported by the USAID Golden Rice
Project. The seed materials were obtained through international collaboration
between IRRI and BRRI (Bangladesh Rice Research Institute). We thank Dr.
N. Baisakh for scientific discussion and Dr. Bill Hardy for editorial
Ahman, E., H. Allen, G. Beaton, B. Benoist, B. Flores, S. Gilespe, S.
Robeneck and F. Viteri, 2000. Nutrition through the life cycle. 4th
Report on the World Nutrition Situation: ACC/SCN in collaboration with
IFPRI, Geneva, Switzerland, p. 23-27.
FAO, 1993. Rice in human nutrition. FAO Food and Nutrition Series No.
26. FAO, Rome.
Goto, F., T. Yoshihara, N. Shigemoto, S. Toki and F. Takaiwa, 1999. Iron
fortification of rice seed by the soybean ferritin gene. Nat. Biotechnol.
Lucca, P., R. Hurrel and I. Potrykus, 2001. Genetic engineering approaches
to improve the bioavailability and the level of iron in rice grains. Theor.
Appl. Genet. 102: 392-397.
Vasconcelos, M., K. Datta, N. Oliva, M. Khalekuzzaman, L. Torrizo, S.
Krishnan, M. Oliveira, F. Goto and S.K. Datta, 2003. Enhanced iron and
zinc accumulation in transgenic rice with the ferritin gene. Plant
Sci. 164: 371-