As the field of biotechnology grows, chemists and biologists face an ever-growing conundrum of ethical obstacles posed by technological breakthroughs. Genetic engineering is on the verge of a revolution. Breakthroughs in genetic engineering (GE) technologies will soon make it easier, cheaper, and more effective than ever to alter the DNA of living organisms. GE technology is used to alter the DNA of living organisms and has many different applications. One of the most promising applications of GMO technologies is gene therapy. Gene therapy is a technique by which biologists alter the DNA of living subjects in an attempt to cure genetic diseases (Gura, 2001). Early controversy over the use of gene therapy to treat the disease was galvanized by the death of a hemophilia patient in a gene therapy trial (Gura, 2001). The biggest obstacle faced by any emerging GE technology is the uncertainty of the risks associated with it. GE's most advanced technology today, CRISPR/Cas9, is no exception to this rule. Although CRISPR/Cas9 is poised to revolutionize the world of genetic engineering, there is considerable resistance from the general scientific community regarding the use of CRISPR/Cas9 and the potential side effects that may unknowingly arise from its use (Gilles & Averof, 2014). A brief overview of DNA/RNA technology, as well as CRISPR/Cas9 technology, combined with an analysis of the potential disadvantages and advantages of CRISPR/Cas9, will allow the reader to better understand the dynamics of the controversy and come to their own consensus on the topic . .Say no to plagiarism. Get a tailor-made essay on "Why Violent Video Games Shouldn't Be Banned"? Get an Original Essay To understand the mechanism of CRISPR/Cas9 you need to be familiar with the function and process of DNA and RNA within the cell. DNA, deoxyribonucleic acid, is the building block of all life. It consists of two chains of complementary nucleotides. Each nucleotide consists of a phosphate group (PO4), a deoxyribose sugar arranged as a 5C ring (C5H10O4), and a nitrogenous base (adenine, guanine, thymine, cytosine). Nucleotides are covalently linked to each other by a phosphodiester bond formed between the 5C sugar of one nucleotide and the phosphate group of another nucleotide. The two strands of nucleotides are linked by hydrogen bonds formed between complementary nitrogenous bases. Adenine has complementarity with thymine, while guanine has complementarity with cytosine (Thieman & Palladino, 2013). The two strands of nucleotides form a double-stranded double helix structure called DNA. DNA is essential for cellular function as it encodes all proteins present in a cell. To produce a protein, DNA must first be copied from another nucleic acid, RNA. The RNA then sends a copy of the DNA to ribosomes in the cell which translate the RNA sequence into a chain of amino acids to form a protein (Thieman & Palladino, 2013). The entire process of transcription and translation is outside the scope of this article, as such it is sufficient to understand the fundamental function of DNA as the “code” for life, and RNA as a translator of DNA. DNA sequences are “read” as letters indicating nitrogenous bases following the direction from 5 primes (5') to 3 primes (3'). 5' refers to the end of the sequence with a phosphate group attached to the fifth carbon of the deoxyribose sugar. For example, a sequence of adenine-thymine-guanine-cytosine would be read as ATGC. A particular DNA sequence with a known function is called a gene. The location of a gene is called a locus. The clustered short palindromic repeatsregularly interspaced proteins, or CRISPR, arise from a natural immune response of bacteria to viral infection (Gilles & Averof, 2014). CRISPR refers to one or more loci present in the genome of bacterial cells. The CRISPR mechanism involves the incorporation of viral DNA into the CRISPR sequence to allow the bacteria to produce an RNA strand complementary to the viral DNA. This is referred to as “CRISPR-derived RNA (Gilles & Averof, 2014)” or crRNA. The crRNA binds to “CRISPR-associated proteins (Cas) to form an active CRISPR/Cas endonuclease complex (Gilles & Averof, 2014).” An endonuclease is a protein that has the ability to degrade DNA. CRISPR/Cas9 refers to a specific endonuclease produced by the bacterium Streptococcus pyogenes. CRISPR/Cas9 contains two forms of RNA and the Cas9 protein. The crRNA contains the sequence necessary to bind complementaryly to the viral DNA. Another type of RNA, “trans-acting antisense RNA, also known as tracRNA,” contains the sequence necessary to “form a complex with Cas9 (Gilles & Averof, 2014).” Together crRNA and tracRNA form the “guide RNA” of the complex. The final piece, Cas9, is a protein that acts as a nuclease in the complex. The CRISPR/Cas9 mechanism requires that a short sequence of nucleotides that follows the sequence targeted by the guide RNA (gRNA) be present in the target DNA. This sequence, called the “protospacer adjacent motif” (PAM), is necessary for Cas9 function (Gilles & Averof, 2014). The CRISPR/Cas9 complex was modified by scientists to contain any desired gRNA sequence, which allows targeting of any gene that contains PAM. In bacteria, the CRISPR process ends with uniform DNA cleavage that occurs a few nucleotides upstream of the PAM. This is an effective method for bacteria to destroy viral DNA. In eukaryotes, however, CRISPR/Cas9 is used to target a gene and insert, remove, or otherwise modify that gene. Engineered CRISPR/Cas9 complexes exploit two types of repair mechanisms used by DNA (Gilles & Averof, 2014). The early process of non-homologous end joining, or NHEJ, does not require a homologous (DNA containing the same genes with potentially different alleles) strand of DNA for repair. In NHEJ the cut ends of DNA are simply brought together and the bonds are reformed. NHEJ can cause deletion of DNA sequences or can introduce (insert) new DNA into the strand during repair (Reis, Hornblower & Tzertzinis, 2014). The other repair mechanism, homology-directed repair, or HDR, requires a homologous strand of DNA to copy a short section of DNA used to repair the broken strand. HDR can be exploited by introducing homologous DNA containing a mutated or normal form of the gene to be repaired. Removing a gene is called knock-out and is usually performed by NHEJ, while inserting a gene is called “knock-in” and can be performed by NHEJ or HDR (Reis, Hornblower & Tzertzinis, 2014).CRISPR / Cas9 confers a number of advantages over other gene editing technologies. First, CRISPR/Cas9 is much simpler than previously implemented gene editing technologies. Two other technologies that work on a similar principle to CRISPR/Cas9, zinc finger nucleases and TALENs (transcription activator-like effector nucleases), are much more technically difficult. Another advantage offered by CRISPR/Cas9 is specificity, as long as the targeted gene has the correct protospacer-adjacent motif, then CRISPR/Cas9 can be configured to target that gene. Finally, an advantage offered by CRISPR/Cas9 over TALENs is that it is not sensitive to methylation. Methylation inhibits the function of TALENs while it appears to have none.