**4. Monitoring of GMOs**

Due to the resentment of the consumers in utilizing GMOs for food and animal feed purposes, many governments have devised policies to give its people freedom over utilization of GMOs. Policies mainly revolve around detection, proper labeling, isolation of propagation area, and tracking of GMOs. International trading requires standardization of procedures and policies related to GMO monitoring and marketing among trading countries. Moreover, in order to limit the entry of approved varieties across the borders of a country, proper monitoring of GMOs is required.

RNA-based genetic engineering technology is becoming more attractive after the approval of white button mushroom for commercialization without stringent testing by the USDA, as this

Due to the advancements in the field of biotechnology and genetic engineering, new varieties are extensively prevailing in the society. Despite their huge potential for human welfare, their commercialization is controversial. Many people perceive all the GMOs to be bad for their health and environment. People who are aware of the mechanism of genetic engineering are concerned about the unintended modifications and their effect on the soil microorganisms [18], plant-microbe interaction [19], and imbalances in the natural biosystems. GMO's controversy mainly revolves around environmental safety [20], human and animal health [21],

Genetically modified organisms produced by genetic engineering or conventional plant breeding are targeted to enhance the desired commercial traits, but GMOs might exhibit unintended traits as well. In the international meeting on "Genetic Basis of Unintended Effects in Modified Plants," biotechnology industry, government, and academia emphasized that no genetic modification is without unintended effects whether conventional breeding or genetic engineering [24]. The source of unintended modifications could be attributed to gene insertions or deletions involving deletion or disruption of endogenous genes and chimeric protein production which perform abnormal function. Genetic engineering approaches involving tissue culturing and in vitro culturing pose the risk of soma clonal modifications arising from the genetic and epigenetic effects of in vitro cultures [25]. Pleiotropic effects may contribute to the unintended modifications if the transgene plays multiple roles or is the part of multiple pathways in an organism leading to the production of potentially harmful secondary metabo-

Biosafety policies involve principles, procedures, and rules devised and adapted for protecting the environment and health of the individuals against potentially harmful metabolites and toxins. Biosafety involves containment of harmful material to avoid unintentional exposure to

Due to the resentment of the consumers in utilizing GMOs for food and animal feed purposes, many governments have devised policies to give its people freedom over utilization of GMOs. Policies mainly revolve around detection, proper labeling, isolation of propagation area, and tracking of GMOs. International trading requires standardization of procedures and policies related to GMO monitoring and marketing among trading countries. Moreover, in order to

technology does not involve the introduction of foreign DNA [17].

216 Applications of RNA-Seq and Omics Strategies - From Microorganisms to Human Health

concerns over interfering with nature [22], and patent issues [23].

toxic agents produced by genetically modified organisms [27].

**3. GMOs and biosafety issues**

lites [26].

**4. Monitoring of GMOs**

The first step in the monitoring of GMO is the detection of transgene in an organism under question. Many methods are being used to detect the genetically modified varieties. GMOs produced by insertion of DNA fragments can be detected by protein-based assays [Enzymelinked immunosorbent assay (ELISA), Western blotting, etc.] or nucleotide-based assays including PCR. PCR-based detection is the most sensitive method which makes use of sequence-specific primers [28]. Due to the abundance of GMOs in the market, it has become very difficult to keep the sequence information of all the transgenes. The advanced highthroughput technologies for GMO detection/monitoring are developed to detect multiple transgenes or related nucleotide components (promoter, enhancer, and terminator) of the cassette [29, 30] in a single experiment. For rapid PCR at atmospheric temperature, various methods have been developed [31]. DNA microarray chips are being developed which contain the probes against all the transgenes present in the commercial varieties [32]. Sampling and hybridization of DNA of a variety under question can detect the presence of any transgene. More efficient, sensitive, and robust methods are required for proper monitoring.

All the above methods are used for the detection of DNA insertion in the transgenic organisms. However, in the RNA-based GMOs, the detection of transgene requires transcriptomic approaches. Transcriptional methods including RT-PCR, gene expression microarray, and RNA-seq can detect all types of GMOs produced through RNA- or DNA-based methods. In transcriptomic approaches, RNA is isolated from the sample and reverse transcribed to produce complementary DNA (cDNA). Due to resemblance in the biochemical properties of RNA and DNA, DNA is often present in the RNA preparations which is eliminated by treating the sample with DNase enzyme. By avoiding this step of DNase treatment, we get both RNA and DNA in the sample. This crude RNA is transcribed and RT-PCR is used for the detection of RNA or DNA of the transgene.
