**2. Developing transgenics: state-of-the-art strategies**

A plant cell is blessed with three major organelles with their own genomes, namely, nucleus, chloroplast, and mitochondria. Of these three, two genomes are routinely manipulated to incorporate new traits in cultivated plants. There are a number of approaches to transfer and introduce genes into the plant genome, depending upon the choice of explant to be used in transformation experiments, for example, *Agrobacterium*-mediated gene transfer, gene gun, agro-infiltration, sonication, and polyethylene glycol treatment. Of these, *Agrobacterium*mediated and gene gun methods are most commonly used approaches to develop transgenic plants. For nuclear transformation, *Agrobacterium* method is more successful than particle bombardment as a more number of transformed shoots can be recovered from the same number of explant.

thereby, chimeric sectors, carrying wild-type or transgenic plastids, appear in leaves of regenerated shoots. Due to phenotypic masking by the transformed cells, both transgenic and wildtype cells in a chimeric shoot look green in color [6], indicating that antibiotic resistance is not cell autonomous. However, both wild-type and transformed sectors are identifiable using green fluorescent protein (GFP). This visual marker allows visual detection of the fluorescing transproteins because they produce green fluorescence upon illumination with blue or

Introductory Chapter: Transgenics—Crops Tailored for Novel Traits

http://dx.doi.org/10.5772/intechopen.81372

Plants on earth synthesize their food by themselves, owing to harbor solar energy conversion and several other chemical reactions in their cells. One of the reactions carried out in plants is carbon fixation during a process, namely, photosynthesis. During photosynthesis Rubisco catalyzes the inefficient carbon fixation, reviewed extensively elsewhere [19, 20]. This raises a question why carbon fixation during photosynthesis is rate limiting. Major reasons are as follows: first, Rubisco's rate of catalysis is much low, and, second, it has to compete with a nonproductive reaction, oxygenation [21], depending upon the relative concentration of

fixation.

5

plants are

plant [28],

carbon dioxide and oxygen, as well as on temperature. Carboxylation results in CO<sup>2</sup>

of gases, the advantages of concentrating carbon dioxide in the chloroplasts of C<sup>3</sup>

Therefore, plant growth and yield can be improved by two ways: (1) by increasing photosyn-

A number of examples are available in the literature, reporting different versions of Rubisco that would improve photosynthesis [22], but considerable success has not been achieved yet.

objectionable [23]. Glycolate catabolic pathway was introduced in chloroplasts for alleviation of photorespiratory losses in *Arabidopsis thaliana*. Photosynthesis is markedly increased in this engineered pathway, thereby widening the applicability of the technique to cereals, for

Developing chlorophyll in the dark, and chloroplasts that are competent for photosynthesis upon exposure to light, is another promising technique that can be implied to improve the photosynthesis in plants [25, 26]. In a study, *chlB* gene from *Pinus thunbergii* was introduced into the plastome of *Nicotiana tabacum* [27]. Transgenic plants when shifted to light from dark in early development of chlorophyll pigments were observed in leaves of transgenic compared to wild-type plants. This helps us to understand the molecular biology of transgenic

Weeds compete with crops for food, thereby lowering the crop yield and affecting farmers. There are two types of herbicides, (1) selective in nature and (2) used before and after emer-

plants appears to be more promising, but due to the leakage

pathway in rice, a C<sup>3</sup>

ultraviolet (UV) light [18, 6].

**3. Transgenics for agricultural traits**

**3.1. Transgenics to improve crop production**

thesis and (2) by reducing photorespiration.

pathway in C<sup>3</sup>

example, wheat and rice, described in detail elsewhere [24].

angiosperms. Another effort is underway to introduce C<sup>4</sup>

gence of plants from seeds, and are specific to leaf morphology.

Introducing C<sup>4</sup>

using various techniques.

**3.2. Transgenics for weed management**

Genetic transformation process involves a number of steps, including selection of a gene that confers resistance to a particular antibiotic for selection and screening purposes, isolation of a trait-encoding gene, choice of promoters and terminating sequences to control the expression of the gene or genes, choice of explant, and an artificial medium to support explant to regenerate into a complete shoot. For selection and screening, usually two types of markers are used: (1) selectable marker and (2) visual marker [10, 12–14]. Selectable marker could be lethal or nonlethal in nature. Nuclear genome transformation is carried out using lethal markers. Regenerated shoots are normally hemi- or heterozygous and need either further purification of transgenome using selection medium or through selfing depending upon the crop used.

The second genome is the chloroplast genome, the plastome that has been modified in a number of plant species, including model, crop, and tree plants. Plastome is a doublestranded DNA molecule of 152 (*Cinnamomum campohra*) – 218 kb (*Pelargonium*) size [15, 16]. Approximately, 120 genes in various plant species are encoded by the plastome [17]. It looks like that most of the ancestral genes have either been lost during evolution or transferred to the nucleus. A mature mesophyll cell contains up to 100 chloroplasts, and each chloroplast contains 100 plastome copies; therefore, the ploidy number of plastome per cell reaches up to 10,000 copies. Furthermore, this number is doubled for genes that are located in inverted repeat regions of the plastome [17].

When transforming a chloroplast, a universal antibiotic cannot be used given that different plants have variable sensitivity to selective agents; therefore, recovery of the transplastomic shoots is dependent on two things: (1) choice of the selective agent to be used and (2) the concentration of the selective agent that allows regeneration and development of shoots from the transformed cells while killing the non-transformed cells. For example, spectinomycin is used to select transformed cells on selection medium from tobacco, lettuce, tomato, potato, cabbage, oil rape seed, and carrot. However, several monocots, including rice and sugarcane, are naturally resistant to spectinomycin; therefore, streptomycin-containing medium was used for carrying out selection for transplastomic lines.

Initially, only few copies of the plastid genome are transformed and maintained under continuous selection pressure. However, stable lines with uniformly transformed genome copies are recovered on selection medium through a repeated cycle of regeneration [6]. During the period, the wild-type and the transgenic plastids and their genomes gradually sort out; thereby, chimeric sectors, carrying wild-type or transgenic plastids, appear in leaves of regenerated shoots. Due to phenotypic masking by the transformed cells, both transgenic and wildtype cells in a chimeric shoot look green in color [6], indicating that antibiotic resistance is not cell autonomous. However, both wild-type and transformed sectors are identifiable using green fluorescent protein (GFP). This visual marker allows visual detection of the fluorescing transproteins because they produce green fluorescence upon illumination with blue or ultraviolet (UV) light [18, 6].
