Biofortification of Tomato

This document discusses the biofortification of tomato and its importance in addressing folate deficiency. It explores the nutritional problem, crops to be addressed, and the metabolic pathway of folate. The document also discusses the engineering strategies used to enhance folate content in crops. The techniques of plant mutagenesis and gene silencing are explained, along with the technologies used in breeding tomatoes and producing transgenic plants.

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BIORFORTIFICATION OF TOMATO
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Biofortification of Tomato
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Biofortification of Tomato
Introduction
The nutritional problem
Folate deficiency causes a variety of health issues such as cancer, neural tube defects,
and homocysteine. Folate is also referred to as vitamin B9 (Price, et al., 2016). It forms a
group of essential vitamins that are involved in the metabolism of body cells. Folate is an
ester or salt of folic acid. It plays an important role in the synthesis of proteins, DNA, and
RNA. Deficiency of folate in the body leads to cell division impairment and accumulation of
toxic metabolites such as homocysteine. Together with vitamin B12, folate plays a significant
role in regulating the synthesis of red blood cells in the body (Moll, & Davis, 2017). Folic
acid is the synthetic form of folate which becomes converted to folate while in the body.
Folate form the most common substance used in fortification and supplementation of food
since it is easily absorbed by the body. Main plants such as animal products, green
vegetables, and fruits, especially tomatoes, are the main source of folate. Most people absorb
folate insufficient amount from their diets to minimize deficiency, but under some
circumstances, the human body may increase the need for the folate. If there is no
supplementation at all folate deficiency occurs. Folate deficiency commonly occurs in
lactating and pregnant women. Also, individuals with serious gastrointestinal tract conditions
and those restricted from certain diets because of weight loss or medical conditions encounter
folate deficiency. Alcohol dependence individuals and older people with an average of
65years of age also experience folate deficiency (Garcia, et al., 2016). The common signs and
symptoms of folate deficiency include muscle weakness, psychological issues such as
confusion, depression, the problem of understanding and judgment, neurological signs
including tingling, feeling of needle, pins, and burning. Other symptoms include dizziness,
pallor and gastrointestinal signs such as vomiting, nausea, weight loss, diarrhea and

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abdominal pain (Koike, et al., 2015) Causes of folate deficiency include low intake of diet
due to restriction by doctors because of certain conditions. Malabsorption because of liver
problems, age, and bariatric surgery. Lactation and pregnancy that increase the demand for
folate due to new tissue growth in fetus and mother.
Crops to be Addressed
Tomato is the main crop to be discussed in the nutritional problem. Tomato and its
products are rich sources of potassium, vitamin c, and less folate. The most critical
phytonutrient in tomato is carotenoids. Other sources of folate include vegetables, especially
green vegetables (Der Straeten, 2017). The low folate content in vegetables and fruits are
consumed after undergoing processing such as freezing, home-cooking, and canning. The
preparation of such products is accompanied by folate loss.
On the other hand, planting of fruits and vegetables under certain conditions may
result in the harvesting of fruits with low folate content. Folate according to analytical
progress it is known to have an inadequate distribution in fruits with many uncertainties as to
their kinetics and mechanism of degradation and this contributes to folate deficiency which is
a global problem (Puthusseri, et al., 2018). The pregnant and lactating mothers take fruits and
vegetables that have a low content of folate. Therefore, there is a need to perform
Biofortification to increase the quality of folate in fruits to curb the global problem.
Main context
How the metabolite addresses the health issue?
Tetrahydrofolate and folates are essential for human health and other organisms. The
folates take part in the reaction of carbon transfer as cofactors in the synthesis of serine,
glycine, purines, thymidylate, and methionine. Folates cannot be synthesized by the body.
Folate deficiency usually causes a range of diseases such as megaloblastic anemia, some
congenital disabilities such as spina bifida, some cancer, and cardiovascular conditions. To

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help curb the folate deficiency, many countries have come with a mechanism to increase the
folate in some foodstuffs. The mechanism involves fortification of grain products and other
foods using synthetic folic acid. An individual taking the correct amount of folate does not
have folate related problems. Enough folic acid needs to be consumed by lactating and
pregnant women to help the fetus from developing congenital deformities of the spine or
brain such as neural defects including anencephaly and spina bifida.
Metabolic pathway of folate
Intake of adequate amount of folate is very crucial in homeostasis and cell division.
Folates usually act as vital coenzymes in several biological pathways such as biosynthesis of
thymidylate and purine (DNA synthesis), metabolism of amino acids, methionine generation,
and DNA methylation (Salway, 2016). Metabolism of folate intersects with the choline
pathway and methionine cycle. The main functions in the process include s-
adenosylmethionine (SAM) formation, which form the vital methyl donor inside the cell and
methylation of Hcy. Most coenzymes of folate are found in the liver.
The enzyme dihydrofolate reductase reduces folic acid to dihydrofolate and then
dihydrofolate to its active form, which is tetrahydrofolate (Ducker, & Rabinowitz, 2017).
The metabolism of cytosolic folate has interrelated cycles usually three in number (The first
cycle begins from 10-formltrihydrofolate and leads to purine production. The other two
cycles utilize 5,10-methylenetetrahydrofolate to form methionine and dTMP.
The predominant form of folate is 5,10methylTHF consisting of 82%-93% of
complete folate in the serum. After the uptake of 5-methylTHF in the cell, it is after that
converted into THF and the reaction is controlled by cobalamin. Methionine cycle is a
pathway necessary for the formation of SAM and also converts Hcy to methionine. The
increased concentration of human serum of the total Hcy can be triggered by vitamin B
deficiency (vitamin B12, B6, and folate) or genetic defects. (Stacey, & Stacey, 2017). In an

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irreversible reaction, 5,10-methylenetetrahydrofolate reductase converts 5,10-methyleneTHF
to 5,methylTHF. Deficiency of vitamin B12 causes a functional deficiency in folate. The MS
is usually in its inactive form after it has been exposed to nitrous oxide or in Vitamin B12
deficiency. Due to the irreversible reaction of the used enzyme MTHFR folate is taken as 5-
methylTHF, and because of that, it cannot be transformed to THF through MS (O'connor,
2016). The synthesis of polyglutamate ceases and prevents intracellular polyglutamate hence
causing a halt in the synthesis of thymidylates and purines.
Engineering strategies
Folate is a molecule with tripartite composing of ᶈ-aminobenzoate, pteridine and
glutamate moieties with a short, ¥-linked additional chain of glutamate attached to the first
glutamate. The biosynthesis of folate in plants is usually highly compartmentalized together
with pteridine released in the PABA and cytosol that are produced in plastids. The moieties
then lead to the formation of dihydropteroate after condensation in the mitochondria that are
later glutamylated, forming folates (Olivares, Aguiar, Rosa, & Canellas, 2015). The modest
rise in folate was mostly associated with depleted pools of PABA in the plant engineering
that suggests the supply of PABA had reduced for the synthesis of folate. The challenge was
currently addressed in the study production of PABA in fruit ripening of tomato and therefore
merging the trait with overproduction of pteridine (Ilahy, et al., 2018). The work was very
significant in demonstrating the feasibility of modifying crops with enough folate to supply
people with the recommended diet. The techniques also shed light to control the pathway of
folate biosynthesis in plants.
In crops, metabolic engineering strategies have been developed using various
approaches. Most of them, including a gain of function using transgenesis and loss-of-
function which is carried out through mutagenesis and gene silencing. There are two
engineering strategies used in quality improvement of crops. They include; Plant mutagenesis

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and gene silencing of specific genes. These engineering strategies can be applied to several
crops or they can all be used to improve the quality of one particular type of plant. The
quality of fruits is enhanced at the cellular level.
Plant mutagenesis
Mutagenesis is the process where there is an occurrence of heritable changes in an
organism's genetic information which is not triggered by either genetic recombination or
segregation but is induced by physical, biological or chemical agents (Oladosu, et al., 2016).
There are three types of mutagenesis that are employed in mutation breeding. They include;
induced mutagenesis where mutations occur due to irradiation such as X-rays, Gamma rays
and ion beam or treatment using chemical mutagens and site-directed mutagenesis that
involve creating a variation at a specific site in the molecule of DNA. Finally, there is
insertion mutagenesis that involves the direct introduction of DNA either through the
transformation of genes or when the transposable elements are activated. Plant breeding,
therefore, need genetic variations of essential traits for improvement of crops (Stoddard, et
al., 2016). However, many mutant genes form the main source of diversity in genes for crop
breeding and functional analysis of the genes of target in most cases. Several Standardized
procedures which induce mutations have been developed for the production of different crops
that improve their quality and commercial interest. Ethyl-methane sulfonate is used in the
chemical procedure and considered simple in achievement. The EMS has demonstrated to be
important inducers of mutagenesis, which is most reliable. EMS aims to generate some
changes randomly in the structure of nucleotides, producing single-nucleotide polymorphism
or deletions, which affect the working of some specific genes in the genome. The main
important aspect in mutation breeding involves individual identification, which has a target
mutation that involves two main steps, including mutant confirmation and mutant screening

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(Lin, et al., 2018). Mutant confirmation is where the putative mutants are re-evaluated under
replicated and controlled environment using a large number of samples. In mutant screening,
individuals that meet selection criteria such as disease resistance and early flowering as
compared to the parent are selected from a population which has undergone mutation.
Gene silencing
Gene silencing is also referred to as post-transcriptional gene silencing or RNA
interference. A specific fragment of the gene in an antisense orientation is inserted in the
crop, making the target RNA to be degraded, therefore diminishing the amount of mRNA and
protein from 45% to 90%. The gene silencing process has been extensively described and is
stated RNA degradation in the cell emerging from antisense RNA hybridization and the sense
of endogenous RNA the specific gene (Poulet, et al., 2017). The target gene triggers the
RNAi pathway. Some fragments are generated, including small interfering RNA which are
about 20-23 nucleotides measured in length that allows systematic degradation of the specific
mRNA through a complex called host RNA-induced silencing complex. Currently, the
siRNA is commonly used in the suppression of the expression of genes and also in the
assessment of gene function. Gene silencing is regarded as a gene knockdown mechanism
where the expression is reduced by 99%. The gene that is responsible for the poor quality of
the fruits is knocked down, enhancing the quality of the tomato.
Technologies used
Breeding of tomato
Techniques of breeding tomatoes have become critical for improving production with
an algorithmic growing in the population and with the existence of the extreme
environmental changes. The development of technologies such as molecular biology together
with bioinformatics has come up with opportunities for an effective plant breeding in fruits.
Breeding objectives have been established to achieve and improve the quality of the

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tomatoes. The yield of fruit is determined by the fruit set efficiency and the cell number and
fruit size. Another common objective of breeding is the resistance of diseases and pests,
which are most destructive and may interfere with the quality of fruits. The release of tomato
genome sequences that form important polygenic resistance exploitation, which only can be
obtained when the responsible molecular markers to the genes meant for resistance are
present. Breeding to improve the quality of tomato entails the physical characteristics
including the shape, color size, and chemical factors such as taste, sensor factors, and acidity.
The objectives of tomato breeding are attained using several breeding techniques such as
mass selection, hybrid development, and pedigree method of tomato breeding.
Transgenic plants productions which articulate a functional gene
Transgenesis involves the insertion of transgenes into a living organism such as a
plant incorporating new traits over the next generations. Intragenic and Cisgenia are used
when the transgenes belong to the related or same plants, respectively (Staskawicz, &
Dahlbeck, 2017). The transgene functionality in a new crop is reached since the genetic code
of all the plants is the same. Therefore, a specific DNA sequence encodes a similar protein in
several plants. The process of transgenesis usually occurs naturally without possible
intervention from humans. Many fruits especially tomatoes have been improved using this
technique
Future perspectives
The methods of improving the quality of tomato from the time they are planted,
especially folate, have been achieved in some cases. Although the common health problem
just been reduced by a small percentage since most of the farmers don’t take in consideration
the need to carry out the genetic engineering for quality improvement of fruits. Others face
some challenges in the field while carrying out the Biofortification. The challenges occur
during the breeding where the genetic variability for the responsible for micronutrients in the

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tomato gene pool and also the required to produce cultivars with the desired characteristics
(Barea, 2015). In some circumstances, the challenge can be curbed by cross-breeding so that
the traits are introgressed into the commercial traits. For Biofortification strategies to be
achieved effectively, the post-harvesting practices of each crop have to be put into
consideration. The practices such as milling and polishing usually consume important
minerals and nutrients in the process and the loss of the required nutrients is dependent on the
genotype. Post-harvesting practices should be minimized in the future to reduce the loss of
essential nutrients that could otherwise cause health-related issues. Also, there are
antinutrients which tend to reduce the availability of essential nutrients in most of the crops
(Filho, et al., 2017). For instance, antinutrients such as tannins, fiber, oxalate, and phytate
tend to reduce the bioavailability of minerals in the gut of a human. It is necessary to reduce
the presence of antinutrients in the food crops before consumption, especially tomato, to
increase the availability of essential nutrients in the human body.

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