**1. Introduction**

Genetic engineering is an advanced field of biology that deals with modification of genomic deoxyribonucleic acid (DNA) in the living organisms to introduce desired traits to benefit mankind. Through genetic engineering, a DNA fragment (gene) is isolated from the donor organism

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and transferred to the recipient where it can be transcribed into messenger RNA (mRNA) and translated to proteins by utilizing the recipient machinery. The donor protein in the recipient system performs its targeted function to modify the desired character of recipient plant, animal, or microorganism. Genomic DNA manipulations may involve addition of a foreign gene from another genome, deletion of an existing gene, or enhancing the expression of an indigenous gene. RNA interfering (RNAi) technology is used to silence the expression of an unwanted gene by inhibiting the mRNA availability for protein synthesis [1]. The genome-level genetic engineering approaches require an insight in the genome, transcriptome, and metabolome [2] of the organisms under study. Like all other applied fields, genetic engineering requires comprehensive information about the genome structure of the donor and recipient before genetic modification. Decision about the morphological character that needs to be improved, the choice of a particular donor and recipient species, genetic networks, and metabolic pathways involved in the expression of a specific trait need to be explored.

transcriptase PCR (DDRT-PCR), gene expression microarray, and next-generation sequencing (NGS) techniques are high-throughput techniques which are currently used. DDRT-PCR can study the expression of hundreds of genes at the same time, whereas microarray and NGS can study the whole transcriptome in a single experiment. Expression microarrays can give insight of the comparative transcriptomics, whereas NGS can provide absolute quantification of each transcript. All these techniques help in the identification of genes which give

Transcriptome Analysis and Genetic Engineering http://dx.doi.org/10.5772/intechopen.69372 215

These identified genes serve as targets to be used in different genetic engineering events. These genes are manipulated in the living organisms to produce GMOs. The modified organism is tested for the proper functioning of the transgene by single gene transcriptional analysis. Then the GMO is tested for the potential risks to the environment and human/animal health using targeted approaches which are biased and require preexisting knowledge of the risk. The comprehensive and unbiased assessment of the GMO should be done using global transcriptome analysis of the GMO with the commercial safe variety. After biosafety testing, GMO is released for commercialization and human/animal utilization. There is great deal of resentment and resistance against utilization of genetically altered organisms. Many governments have designed policies to properly monitor the presence and movement of GMOs. Transcriptional analysis is widely being utilized for the monitoring of various newly devel-

Genetic engineering is the field of science which is revolutionizing the world by manipulating the genome and transcriptome of living organisms to introduce desired traits in them. Since the commercialization of "Flavr Savr" tomato in 1994 [10], 357 GM crops belonging to 27 species all over the world have been commercialized [11], and this number is increasing day by day. Genetic engineering is widely being used for the improvement of crops, animals, fungi [12], bacteria [13], and other organisms to benefit mankind. Insect resistance, herbicide resistance, disease resistance, and abiotic resistance are being incorporated in the industrially important crops to make them tolerant to stresses. Yield and nutritional content of food crops are being modified to improve the feed for humans and animals. Scientists [14] produced transgenic maize with overexpressing *Oryza sativa* myeloblastosis 55 (OsMYB55) gene and found that the transgenic maize became more tolerant to heat and drought stress through activating the expression of stress-responsive genes. Microorganisms (bacteria and fungi) are being genetically engineered for the production of useful enzymes [13], secondary metabolites, beneficial oils [12], and antibiotics on commercial scale to be utilized in the pharmaceuti-

In 2010, 29 countries were growing genetically modified crops, and 31 countries had the approval to import GM crops. In USA, more than 94% of the cultivated soybean and cotton while 92% of corn is genetically modified [15]. The commercialization of the first genetically modified animal "AquAdvantage Salmon" for food was approved recently in 2015 [16].

differential expression under different conditions.

**2. Genetic engineering for human benefit**

oped organisms.

cal, food, and medical industry.

Transcriptome analysis is a robust and cost-efficient method which provides information about the internal biological processes, cellular biosynthesis, and metabolic functions of a cell, tissue, or living organism [3]. This technique can be utilized by the genetic engineering scientists for the identification and quantification of genetic factors which positively or negatively regulate a particular trait of interest [4]. Comparison of gene expression profiles of an organism exhibiting the desired traits with the genetically similar organism lacking that trait can help in the identification of genetic factors involved in the development of that trait [5, 6]. These genetic factors might affect that trait positively or negatively. Enhanced accumulation of a particular transcript in the organism with desired phenotype as compared to the reference organism indicates that overexpression of that transcript is required for the exhibition of that trait. This phenomenon is called as positive regulation. In negative regulation, reduced expression of a gene is responsible for the exhibition of a desired trait [7, 8].

Positively regulated genes serve as genetic engineering tools for overexpression of a gene regulating a particular trait resulting in the introduction of that trait in genetically modified organism (GMO). For example, in transgenic cotton, expression of crystal protein (Cry10Aa) is responsible for resistance against boll weevil [9]. Advances in gene silencing technology through RNAi have led to utilization of genes which are negatively correlated with the desired traits. In cotton plant, seed-oil content increased by 16.7% by silencing GhPEPC1 gene through RNAi technology [8].

Transcriptome analysis and genetic engineering go hand in hand in the modern era of genetic improvements. Comparative transcriptional studies using single gene approaches or high-throughput approaches are used to identify the differentially expressed genes in a specific condition/organism as compared to reference. In single gene approaches, the expression of a gene of interest is quantified in different sets of conditions/tissues using northern blotting or reverse transcriptase polymerase chain reaction (RT-PCR). Northern blotting technique utilizes the gene-specific probes for comparative quantification of mRNAs of the target gene, whereas RT-PCR uses gene-specific primers to amplify and subsequently quantify the mRNA molecules. High-throughput technologies have the power to measure and analyze the expression of all the genes in a set of conditions. Differential display reverse transcriptase PCR (DDRT-PCR), gene expression microarray, and next-generation sequencing (NGS) techniques are high-throughput techniques which are currently used. DDRT-PCR can study the expression of hundreds of genes at the same time, whereas microarray and NGS can study the whole transcriptome in a single experiment. Expression microarrays can give insight of the comparative transcriptomics, whereas NGS can provide absolute quantification of each transcript. All these techniques help in the identification of genes which give differential expression under different conditions.

These identified genes serve as targets to be used in different genetic engineering events. These genes are manipulated in the living organisms to produce GMOs. The modified organism is tested for the proper functioning of the transgene by single gene transcriptional analysis. Then the GMO is tested for the potential risks to the environment and human/animal health using targeted approaches which are biased and require preexisting knowledge of the risk. The comprehensive and unbiased assessment of the GMO should be done using global transcriptome analysis of the GMO with the commercial safe variety. After biosafety testing, GMO is released for commercialization and human/animal utilization. There is great deal of resentment and resistance against utilization of genetically altered organisms. Many governments have designed policies to properly monitor the presence and movement of GMOs. Transcriptional analysis is widely being utilized for the monitoring of various newly developed organisms.
