Skip to main content

TRANSGENIC RICE - Review (AK Gupta, Ph.D)

Posted in



Micronutrient induced malnutrition is widely prevalent in Asia, approximately 24 thousand people die from malnutrition every day. Statistics shows that despite of enormous efforts and investments in conventional breeding, we have developed so many varieties which are high yielding and good in nutrition quality, but we are still facing vitamin A deficiency affects more than 400 million people worldwide, leaving them vulnerable to infection and blindness. Iron deficiency affects 3.7 billion people particularly women leading to high maternal deaths and infant mortality. Developing micronutrient dense rice, with higher amount of iron, zinc and vitamin A, can have a beneficial impact or the health of low-income people. Conventional breeding when combined with biotechnology can provide powerful tools to achieve this goal. Ye et al. (2000) produced transgenic rice (golden rice) with high source of provitamin-A (beta-carotene) through biosynthetic pathway engineered into its endosperm by using agro-bacterium mediated transformation to introduce three genes, phytoene synthase, lycopean cyclase and phytoene desaturase from Daffodil plant (provided 2 genes) and Erwinia uredovora (provided one gene), respectively. Iron content in rice can be increased upto two-fold by introducing a ferritin gene from Phaseolus vulgaris into rice grains and its bioavailability may be increase by introduction of a thermo tolerant phytase gene from Aspergillus fumigatus (Lucca et al., 2002). The expected outputs of the colloquium is assessment of role of molecular breeding in the nutritional environment of rice and developing strategies for overcoming hidden hunger caused by micronutrient deficiencies through an integrated strategy involving the use of biofortifieds rice and underutilized crops like millets and grain legumes. In this article, we have reviewed the application of biotechnology tools and their impact towards improving the nutrition quality in rice.

The problems

The major problem is that, to date approximately 840 million people surfer from all malnutrition and every day 24,000 die from malnutrition. This situation become more severe in the form 250-500 thousand children became blind each year from vitamin A deficiency. The major cause is that 1.3 billion people affected by poverty based lack of sufficient nutritious food, in fact these people are not capable to purchase fruits and vegetables which are very rich source of micronutrient, protein and energy and they are totally depend on grain food for their nutrition requirement (James, 2001). These problems can be solve by traditional intervention as like supplementation is useful for producing a rapid improvement in Fe status in anemic individuals, but is expensive and usually has poor compliance because of the unpleasant side effects of medicinal iron. In India, vitamin A supplementation program has been in operation since early 1970, (Reddy, 1994). However, national coverage of all the children has been hard to sustain over time. Artificial food fortification has been considered the best long-term strategy for prevention, but there are technical problems related to the choice of a suitable iron compound. The iron compounds of relatively high iron availability, such as ferrous sulfate, often provoke unacceptable colour and flavour changes, whereas, those compounds which are organoliptically inert, such as elemental iron, are usually poorly absorbed (Wijk, 2001).

Diet diversification is undoubtedly the most logical and sustainable strategy to improve vitamin A status. It tradionally involves the attempt to increase the consumption of grain, vegetables and suitable fresh fruits. But this approach is more complex, involving a number of factors including accessibility, affordability, and change in dietary habits. An alternative more sustainable approach would be the enhancement of micronutrient of the good staples by plant breeding. But it will take a very long time to achieve the goal. So keeping the view the limitations of the additional interventions measure, it is desirable to deliver the micronutrients by sustainable basis, through such a vehicle that is a staple food of the target population. But unfortunately, rice which is the good staple for more than half of the world population does not accumulate Pro-vitamin A in its endosperm (Tan et al, 2004).

In fact over dependence on rice for food is considered to be the major cause for such micronutrient deficiencies. However, by using genetic engineering to bioengineer  carotene synthesis pathway into rice endosperm (Ye et al, 2000) and by introducing genes like Ferritine, Phytase from soybean and Aspergillus fumigatus respectively, into the rice endosperm is ultimate tool to provide nutrient dense food. During the nine year period 1996-2004, the global area of GM crops increased more than 47 fold from 1.7 million ha to 81.0 million ha in 1996 and 2004 respectively. Even, china has projected potential grain of $ 5 billion in 2010, $1.0 billion from Bt cotton and $ 4 billion from Bt rice (James, 2004). This higher rate of Biotech crops adoption by farmers itself reflects the scope of genetically engineered crops and promote scientist to use such technology.

What are transgenic plants?

A transgenic crop plant contains a gene or genes, which have been artificially inserted, instead of the plant acquiring them through pollination. The inserted gene sequence may come from another unrelated plant, or from completely different species; transgenic Bt corn, for example, which produces its own insecticide, contains a gene from a bacterium. Plants containing transgenes are often called genetically modified or GM crops, although in reality all crops have genetically modified from their original wild state by domestication, selection and controlled breeding over long periods of time.

Why make transgenic crop plants?

A plant breeder always tries to assemble a combination of genes in a crop plant, which will make it as more useful and productive as possible. Depending on where and for what purpose the plant is grown, desirable genes may provide features such as higher yield, pest or disease resistance or tolerance to heat, cold and drought. Combining the best genes in one plant is a long and difficult process, especially as traditional plant breeding has been limited to artificially crossing plants within the same species or with closely related species to bring different genes together. e.g. Gene for protein in soybean could not be transferred to a completely different crop such as corn using traditional techniques. Transgenic technology enables plant breeders to bring together in one plant useful genes from a wide range of living sources, not just from within the crop species or from closely related plants. This technology provides the means for identifying and isolating genes controlling specific characteristics in one kind of organism, and for moving copies of those genes into another quite different organism, which will then also have those characteristics. This powerful tool enables plant breeders to do what they have always done-generate more useful and productive crop varieties containing new combinations of genes-but it expands the possibilities beyond the limitations imposed by traditional cross-pollination and selection techniques.

How do you make a transgenic plant?

Strategies to make a transgenic plant we can divide in following subheads:

Introduction to DNA

Locating gene for plant traits

Designing genes for insertion


Selection and regeneration


1. Introduction to DNA

The underlying reason that transgenic plants can be constructed is the universal presence of DNA in the cells of all living organisms. This molecule stores the organism’s genetic information and orchestrates the metabolic processes of life. Genetic information is specified by the sequence of four chemical bases (adenine, cytosine, guanine and thymine) alone the length of the DNA molecule. Genes are discrete segments of DNA that encode the information necessary for assembly of a specific protein. The proteins then function as enzymes to catalyze biochemical reactions, or as structural or storage units of a cell, to contribute to expression of a plant trait. The general sequence of events by which the information encoded in DNA is expressed in the form of proteins via an mRNA intermediary is shown in the diagram below.

Transcription Translation

DNA mRNA Protein trait

2. Locating genes for plant traits

Identifying and locating genes for agriculturally important traits is currently the most limiting step in the transgenic process. We still know relatively little about the specific genes required to enhance yield potential, improve stress tolerance, modify chemical properties of the harvested product, or otherwise affect plant characters. Usually, identifying a single gene involved with a trait is not sufficient, we must understand how the gene is regulated, what other effects it might have on the plant, and how it interacts with other genes active in the same biochemical pathway. Public and private research programs are investing heavily into new technologies to rapidly sequence and determine functions of genes of the most important crop species. These efforts should result in identification of a large number of genes potentially useful for producing transgenic varieties.

3. Designing genes for insertion

Once a gene has been isolated and cloned, it must undergo several modifications before it can be effectively inserted into a plant.

Marker gene Promoter Transgene Terminator sequence


Simplified representation of a constructed transgene, containing necessary components for successful integration and expression.

  1. A promoter sequence must be added for the gene to be correctly expressed. The promoter is the on/off switch that controls when and where in the plant the gene will be expressed. To date, most promoters in transgenic crop varieties have been constitutive i.e., causing genes expression throughout the life cycle of the plant in most tissues. The most commonly used constitutive promoter is CaMV35S, from the cauliflower mosaic virus, which generally results in a high degree of expression in plants. Other promoters are more specific and respond to cues in the plant’s internal or external environment. An example for light-inducible promoter is the promoter from the cab gene, encoding the major chlorophyll a/b binding protein.

  2. Sometimes, the cloned gene is modified to achieve greater expression in a plant. For example, the Bt gene for insect resistance is of bacterial origin and has a higher percentage of A-T nucleotide pairs compared to plants, which prefer G-C nucleotide pairs. In a clever modification, researchers substituted A-T nucleotides with G-C nucleotides in the Bt gene without significantly changing the amino acid sequence. The result was enhanced production of the gene product in plant cells.

  3. The termination sequence signals to the cellular machinery that the end of the gene sequence has been reached.

  4. A selectable marker gene is added to the gene ‘construct’ in order to identify plant cells or tissues that have successfully integrated the transgene. This is necessary because achieving incorporation and expression of transgenes in plant cells is a rare event, occurring in just a few percent of the targeted tissues or cells. Selectable marker genes encode proteins that provide resistance to agents that are normally toxic to plants, such as antibiotics or herbicides. As explained below, only plant cells that have integrated the selectable marker gene will survive when grown on a medium containing the appropriate antibiotic or herbicide. As for other inserted genes, marker genes also require promoter and termination sequences for proper function.

4. Transforming plants

Transformation is the heritable change in a cell or organism brought about by the uptake and establishment of introduced DNA. There are two main methods of transforming plant cells and tissues.

(a) The “gene gun” method (also known as microporjectile bombardment or biolistics) for the transformation of regenerable tissue (Sanford and Klein, 1987, Christou et al, 1998) offered an easier way to transform recalcitrant higher plants. It is the introduction of substances into intact cells and tissues through the use of high velocity microprojectile via a mechanism that breaches cell walls and membranes (Christou, 1997) A microprojectile should be small enough to enter a cell a tissue in a non-lethal manner, and should be capable of carrying DNA on its surface or in its interior. Typically these are made of high density metals such gold or tungsten, which are more or less spherical and approximately 1.5 – 3.0 m fold particle are used in rice transformation in the laboratory at IRRI (Datta et al, 1997).

Typically DNA is loaded onto 1.5 – 3.0 m gold bead at a rate of upto 40g DNA/mg of gold, using cacl2 and supermidine (Klein et al, 1987) to precipitate the DNA to the gold and particle delivery system PDS-1000/He uses a shock wave generated by the sudden release of compressed helium to accelerate a thin plastic sheet into a metal followed by ethanol treatment.

Various genes of agronomic importance have been introduced in rice by different groups, including Bt endotoxin (Alam et al, 1996; 1998; Datta et al, 1998; Nayak et al, 1997; Tu et al, 1998a; Wu et al, 1997), bar gene (Datta et al, 1992; Cao et al, 1992; Ho et al, 2001; Kaur, 2002) Xa 21 (Tang et al, 1999; Nagadhara et al, 2003), chitinase gene for sheath blight resistance (Lin et al, 1999; Baisakh et al, 1999;2001; Datta et al, 2001), phtoene synthase gene (Burkhardt, et al, 1997), ferritin gene (Drakakaki et al , 2000; Vasconcelos et al, 2003). But it is limited to the copy number and rearrangement of the introduced DNA is high become prone to gene silencing (Oard et al, 1996) and Ca using genomic changes. However, to date it is still considered as a very efficient tool for production of phenotypically normal, fertile transgenic japonica and indica rice cultivars (Jain et al, 1996).

(b) The agrobacterium method : Transformation via agrobacterium has been successfully practiced in dicots (broadleaf plant like soybeans and tomatoes) for many years, but only recently has it been effective in monocots (grasses and their relatives). In general, the agrobacterium method is considered preferable to the gene gun, because of the greater frequency of single-site insertions of the foreign DNA, making it easier to monitor. Agrobacterium tumefaciens is a gram negative soil bacterium that can genetically transform plant cells by transferring a define piece of DNA (Known as T-DNA) from its tumour-inducing (T1) plasmid into the genome a infected plants results in crown gall tumors, the tumour provides substances for Agrobacterium tumefacioens to grow on the plate with a set of 25 expressible vir genes (……………..) into seven operons within the Ti-plasmid. Other genetic elements are Agrobacterium chromosomal gene (Chv) and right border and left border of T-DNA are necessary to constitute the T-DNA transfer machinery. Hiei et al, 1994 gave an efficient transformation protocol for japonica cultivars and recommended that the three week old scutella-derived embrogenic calli are the most suitable material for infection by Agrobactrium. They cultivated calli with (BA 4404 (pTOK 233, pIG121Hm) and EHA 101(pTOK 233, pIG121Hm) for three days in the presence of 100..m acetosyringone, the transformation efficiency was 23%.

To date, several Ti plasmid based vector system are available for plant transformation. Typically binary vector comprising of octapine type vir helper strain such as LBA 4404 (Hoekema et al., 1983) that harbors the disarmed Ach Ti plasimid and a binary vector such as pBin 19 is very commonly used for transformation. Agrobacterium-mediated transformation system has been reported by different groups for japonica (Chen et al.,1993; Hiei et al.,1994; Komari et al.,1996; Toki et al., 1997; Cheng et al., 1998; Yokoi et al,… and Ye et al., 2000) and indica rice varieties (Aldemita and Hodges, 1996; Datta et al., 1996; Rasid et al., 1996; Baisakh et al., 1999; Datta et al., 2000; Azakanandam, 1999; Datta et al., 2003). Application of this method concise its area only to the tissue culture responding genotypes.


5. Selection and regeneration

Selection of successfully transformed tissues: Following the gene insertion process, plant tissues are transferred to a selective medium containing an antibiotic or herbicide, depending on which selectable marker was used. The hygromycin phophotransferase (hpt) gene has been widely used as a selectable marker gene in Agrobacterium transformation experiments (Hiei et al, 1994; Ku et al, 1999, Cheng et al, 1998 and Ye et al, 2000). Several transformation containing selectable marker gene (65,67). Several strategies for production of marker free transgenic plants have been including co-transformation; a site specific recombination system, and intra genomic relocation of transgenes via transposable elements.


Extraction of plasmid DNA

To obtain plasmid DNA of bacteria, we start from a known quantity of bacterial culture that is centrifuged for 2 min. at 5000 g. The residue is put in alkaline lysis solution in the cold that allows the rapture of cell membranes and consequently the restriction of cells. The cell debris, the ruptured membrane, and the genomic DNA that is linked to these membranes are eliminated by a second centrifugation. A solution of phenol, chloroform and isoamyl alcohol is used to eliminate the proteins. Two more rinses are done in the same conditions of the tube obtained after the second centrifugation is eliminated, while the lower phase, which contains the DNA and RNA, is precipitated by the addition of isopropanol.

A treatment with RNAse at ambient temperature allows us to conserve only the plasmid DNA in the tube.

Vitamin A deficiency

The problem: Problem is that, rice is major staple food for more than 50% of the world’s population, does not contain any provitamin A, which is one of the most important micronutrient for maintenance of human health involved in several critical function of the body such as stimulation of growth, proper development of skeletal tissue, normal reproduction, and preservation of epithelial tissue. In addition to it implicated in quenching of free radicals and preventing oxidative damage as well as in supporting the immune system (Bendieh, 1989 and 1993). It also delay symptoms of HIV (Santamaria and Biachi-Santamaria, 1993; Tang et al 1993; Sembe et al 1995a) and protect from certain types of cancer (Olson, 1992; Ziegher, 1993).

Consequences: Each year more than two million VAD (vitamin A-deficiency) associated childhood deaths and 5 lacks children become blind.

According to the World Health Organization, as many as 230 million children are at risk of clinical or sub clinical VAD, a condition which is largely preventable. VAD makes children especially vulnerable to infections and worsens the course of many infections. According to an estimate approximately 400 million rice-eating people suffer from vitamin A-deficiency, all over the world.

The transgenic concept

Introduce a desirable gene under endosperm-specific regulation of produce vitamin A rich rice.

Biochemical pathway engineering from GGPP

Initial research confirmed that rice endosperm was capable of synthesizing geranyl-geranyl diphosphate, an early precursor of -carotene in the rice endosperm required adding four enzymatic steps sequences of carotenoid biosynthesis genes in Erwinia uredovora and elucidated the carotenoid biosynthesis pathway by functional expression of gene products expressed in E. cali, i.e. Geranyl geranyl pyprophosphate CrtB prephytone pyrophosphate CrtE phytoene Crt1 Lycopene CrtY carotene CrtZ Zeaxanthin Crtx Zeaxanthin-- diglucoside (misawa et al, 1990). However, a gene CrtI alone capable to perform four sequential desaturation steps to convert phytoene to Lycopene.

Burkhardt et al, 1999 demonstrated that engineer -carotene biosynthetic pathway in a non photosynthetic carotenoid lacking plant tissue by transforming japonica rice variety T309 with daffodil phytoene synthase gene driven by either CaMV 355 promoter or glucelin (Gt1) seed specific promoter. The transgenic rice plants accumulated phytoene in the endosperm and higher amount were 0.74 g.

After the review of this experiment Ye et al, 2000 introduced three genes, a daffodoil phytoene synthease (PSY) gene under the control of seed specific Gt1 promoter, an E. Uredvora phytoene desaturase (Crt 1) gene controlled by CaMV 35 promotes, and daffodil lycopene cyclase (lcy) into a japonica rice lines, which resulted in the accumulation of carotenoids in the endosperm upto 1.6 g/g. Man behind the golden rice Potrykus and Beyer chose the daffodil plant as the source of genes coding for the four enzymes. The first task was to introduce the daffodil gene for phytoene synthase under the control of a rice promoter that would ensure expression of the gene only in the rice endosperm. This was done successfully. The engineered rice plants produced phytoene in he endosperm at levels that would, if converted to -carotene, be nutritionally significant. They then set to work to introduce daffodil for the three remaining enzymatic steps required to convert phytoene to -carotene, one of these daffodil genes. However, turned out to be unusually complex and difficult to work with, so they also tried a bacterial gene (Erwinia), coding for an enzyme that could catalyze two steps in the pathway, including the step that was causing the problem. Although they initially intended to introduce the genes independently and then combine them by crossing. They also tried adding them together in one sophisticated transformation experiment.


Phytoene synthase (PSY)


Phytoene desaturase (Crt 1)


Lycopene cyclase (Lcy)

Carotene Carotene

As shown in Fig., the resulting rice plants containing two daffodil gene and one Erwinia gene carried out all four steps in the pathway and produced -carotene in the endosperm. The plants were normal, the only difference being that, after milling, their grains was a beautiful golden yellow. This technology further extends with to several widely grown to indica rice varieties by Datta et al, 2003 and Variety IR 64 (Datta et al, 2000; Hoa et al, 2003).

How much golden rice has a child to eat to defeat vitamin A deficiency?


FAO recommended 0.3 mg/day vitamin for 1-3 years old child so the average amount needed to prevent deficiency state is only 0.15 mg/day.

100 g golden rice contain 0.16 mg -carotene, stored in lipid membranes but the bioavailability conversion factor of -carotene is 2 : 1. So 100 g of golden rice may provide 50% of the total vitamin A required.

Iron deficiency anemia

Iron deficiency is estimated to effect 30% of the world population making iron by for the most widespread nutrient deficiency worldwide (WHO, 1992). The functional effects of iron deficiency result both from a reduction in circulating hemoglobin and in the iron-containing enzymes and myoglobin. The amount of bioavailable iron is dependent both on the iron intake and absorption. Dietary iron in developing countries consists primarily of non-heme iron, whose poor absorption is considered a major factor in the etiology of iron deficiency anemia (Ballot at al., 1987) grain and legume staples are high in phytic acid, which is potent inhibitor of iron absorption.

The problems

As rice is the staple food, does contain a little iron (Fe 0.2 mg-2.8 mg/100 g rice), an inhibitor of iron resorption (phytic acid) and does not support from a vegetable and fruits (Ballot et al., 1987) or muscle tissue (Taylor et al., 1986) is often limited.

The consequences

The major consequences are reduced psychomotor and mental development in infants (Walter et al., 1986), poor pregnancy outcome (Murphy et al., 1986), decreased immense function (Murakawa et al., 1987) and triteness and poor work performance (Basta et al., 1979). In terms of rice, rice-eating poor suffer from iron deficiency and affects nearly 3.0 billion people. In infants and young children even mild anemia can impair intellectual development. Anemia in pregnancy is an important cause of material mortality, increasing the risk of hemorrhage and sepsis during childbirth. Infants born to anemic mothers often suffer from low birth weight and anemia themselves. An inadequate dietary iron intake is the main cause of IDA. According to UNICEF, nearly tow billion people are estimated to be anemic and about double that number, or 3.7 billion are iron deficient, the vast majority of them women. In Africa and Asia UNICEF estimates that IDA contributes to approximately 20 per cent of all maternal deaths.

The transgenic concepts

By using the transgenic technology, we can decrease the inhibitor by introduce the phytase enzyme, increase iron content by introducing a transgene ferritin and can increase the bioavailability by introduce the add uptake-promoting substance like cystein.

Procedure to increase iron content in rice

Lucca et al, 2002 first introduced a ferritin gene from phaseolus valgaris into rice grain, increasing their iron content up two fold. Japonica rice variety Taipaei 309 was used (…………………..) inoculated embryogenic calli, deriven from mature zygotic embryos with agrobacterium tumefaciens strain LBA 4404 containing the ferritin of the metallothionin like …………. gene.

They constructs pAGt lFe containing the gene for the ferritin protein from phascolus vulgaris and pAGt 1Me with metallothirnein like protein followed by agrobacterium-mediated transformation of mature rice embryos and 40 hygromycin resistant clones were obtained so in order to increase iron content, we first introduced a ferritin gene from Phaseolus vulgaris in to rice grains, increasing their iron content upto two fold. To increase iron bioavailability, we introduce a thermo-tolerant phytase from Aspergellus fumigates into the rice endosperm. In addition, as cystein peptides are considered major enhancers of iron absorption, we over expressed the endogenous cystein-rich metallothionin like protein.

This content of cysteine residue increased about sevenfold and the phytase level in the grains about 130 fold, giving a phytase activity sufficient to completely degrade phytic acid in a simulated digestion experiment. So finally they got. This rice, with higher iron content, rich in phytase and cystein-peptide has a great potential to substantially improve iron nutrition.

High iron rice : Increase in iron content

Result of Lucca et al, 2000 showed that TR line is content rgMT gene and WT lines are wild type, which contain from ranged 9.95 to 10.65 mg/g seed. While transgenic line (1-8) contain iron varied from 11.53 to 22-07 mg/g seed. So, it can be estimate that a two fold increase in iron content.

Protein deficiency

Very little work has been done towards increase the protein contents in rice by using transgenic technology. Although their are many examples available to increase protein content by using traditional breeding in crops like wheat and maize.

A rice diet provides less than 20% of the required essential amino acids (Isoleucine, Lucine, Lysine, Thryonine, Tryptophan and Valine). But it contain incomplete profile of sulfur containing amino acids like cystein.

The consequences

Rice-eating poor people are deficient in essential amino acids that impair numerous cellular functions and normal development.

The transgenic concept

Transfer the genetic information for accumulation of a balance mixture of the essential amino acids in the endosperm.


Interventions applied, so far, to reduce both IDA and VAD are (a) supplementation (e.g. distribution of vitamin A capsules), (b) food fortification (e.g. adding iron to wheat flower), and (c) dietary education and diversification. In a FAO/Who World Declaration on Nutrition (1992) the following strategy ahs been advocated : Ensure that sustainable food-based strategies are given first priority particularly for populations deficient in vitamin A and iron, favoring locally available foods and taking into account local food habits. Supplementation should be progressively phased out as soon as micronutrient-rich food based strategies enable adequate consumption of micronutrients. As per Pinstrup-Andersen, Director General of the International Food Policy Research Institute has pointed out that a sustainable solution of the problem will come only when it will be possible to improve the content of the missing micronutrients in the major staple crops. This was exactly what we were trying to achieve. As the necessary genes for such an improvement were not available in the rice gene pool, genetic engineering was the only technical possibility. As rice endosperm did not contain any provitamin A, the task was to introduce the entire biochemical pathway. As rice endosperm contains very little iron and considerable amounts of a potent inhibitor of iron resorption, and as resorption from a vegetarian diet is generally poor, the task was to increase the iron content, reduce the inhibitor content, and add a resorption-enhancing factor. This manuscript discussed methods of transgenic technology towards improving the nutrition quality in rice. Despite the considerable uncertainty that exists about the effect of transgenic crops, a few conclusions can be drawn.

The content and bioavailability of micronutrients in rice is low and under genetic control and can be increased by conventional as well as transgenic technology.

  • When the necessary genetic variability is not available, genetic engineering offers possibilities for increasing the content and bioavailability of micronutrients in crop plants.

  • Biotechnology approaches to manipulate ferritin expression in the rice seed may contribute to a sustainable solution to global problems of iron deficiency.

Finally we can conclude that the advancements made with transgenic plants will continue to have a great impact on human diet. It offers de nova approaches in producing cultivars with high nutrition quality. The progression of transgenic technology new has allowed for the progression of human life and other medicinal advancement.

Your rating: None

Please note that this is the opinion of the author and is Not Certified by ICAR or any of its authorised agents.