A New Dawn of Gene Editing
For millions of years, the CRE recombinase systems have ruled over genome editing. Although they were awesome leaders, a new force of power and greatness emerged from the shadows. The DRE-ROX system was here, and they were not leaving without a fight. Keep reading to witness the showdown between CRE and DRE, as well as the other types of genome editing that sided with CRE and DRE.
Woah Meera that seems so cool! I cannot wait to learn more about the CRE and DRE-ROX systems!
Slow your roll their cowboy. Before we witness the crazy fight between the CRE and DRE-ROX systems, let us brush up on the basics of gene editing, sequencing, epigenetics, and CRISPR.
(if you are already a bio-nerd like me, feel free to skip this section and head right over to the CRE systems!)
The Basics of Gene Editing Refresher
Before we dig too deep into sequencing, epigenetics, CRISPR, and an awesome battle between CRE and DRE, let us refresh our memory on the basics.
- DNA stands for deoxyribonucleic acid. It’s made up of units of biological building blocks called nucleotides. DNA is a vitally important molecule for not only humans but most other organisms as well. DNA contains our hereditary material and our genes — it’s what makes us unique.
- Each stand is coded with a four-letter alphabet: A, T, C, and G. These letters form complementary base-pairs: A only bonds with T, C only bonds with G.
- RNA. One of RNA’s main functions is to copy DNA’s code. RNA’s abbreviation is ribonucleic acid. It is a complex compound of high molecular weight that functions in cellular protein synthesis and replaces DNA (deoxyribonucleic acid) as a carrier of genetic codes in some viruses. The nitrogenous bases in RNA are adenine, guanine, cytosine, and uracil, which replace the thymine in DNA.
- Mutations occur when the order of the genetic code is changed. Gene editing is essentially artificially inducing and specifying a mutation.
Sequencing
Genomic sequencing is a process for analyzing a sample of DNA taken from your blood. In the lab technicians extract DNA and prepare it for sequencing. Within every normal cell are 23 different pairs of chromosomes. Chromosomes are structuring that house DNA. DNA, or Deoxyribonucleic acid, is coiled into a shape called the double helix. The double helix can be unwound into a ladder shape. This “ladder” is made out of paired chemical letters called bases. Overall, our DNA contains about 6 billion bases! In the DNA alphabet, there are four bases called A, T, C, G. In these pairs, A and T only join together, and C and G only join with each other. To read the sequence of bases in DNA samples are inserted into a sequencing instrument where high-frequency sound waves break the DNA into smaller pieces that are only about 600 bases long. Special tags are added to the ends of the fragmented DNA. These tagged strands of DNA can then attach to a glass slide in a sequencer. Each piece of DNA has been copied hundreds of thousands of times which in turn creates clusters of identical DNA fragments. Next, the sequencer reads the DNA one base at a time using different colored tags for each DNA base special sensor within the machine to detect the different colored tags. This sequence of colors reveals the DNA sequence of each fragment. Powerful computers piece together these individual DNA fragments and reveal the sequence of your DNA then medical experts use specialized software to analyze and compare the DNA sequences. This helps identify the genetic variants and completes the process of sequencing.
Epigenetics
Epigenetics is the study of how genetics interacts with the multitude of smaller molecules found within cells that can activate and deactivate genes.
Let us break that sentence down a bit.
Try thinking of DNA as a genetic cookbook.
The molecules are the main chef, they get to decide what gets cooked and when. They are not making any “conscious” choices themselves but instead, their presence and concentration within cells make the difference. So how does this process really work? Genes in DNA are expressed when they are read and transcribed into RNA, which is then translated into proteins by structures inside the cell called ribosomes. Proteins are the main factor in determines the characteristics and functions of a cell.
Epigenetic changes can boost or interfere with the transcription of certain genes. The most common way this interference happens is that DNA, or the proteins it is wrapped around, gets labeled with small chemical tags. The set of the chemical tags that are attached to the genome is called the epigenome. Some of these, like a methyl group, inhibit gene expression by derailing the cellular transcription machinery or causing the DNA to coil more tightly, causing it to be inaccessible. The gene is still there, just silent. Boosting transcription is essentially the opposite. Some chemical tags will unwind the DNA, making it easier to transcribe, which in turn speeds up the process of creating associated proteins. Epigenetics changes can survive cell division, which means the effects are permanent. This however does not mean it is a bad thing, it can be a good thing as well. Epigenetic changes are a part of normal development. The cells in an embryo will start with one master genome. As the cells divide, some genes are activated, while others are inhibited. Over time through epigenetic reprogramming, these cells may turn into heart cells, muscle cells, live cells, and so on.
CRISPR
In my mind, I like to think of CRISPR with an analogy. For example, take a working document. In a document, if I suspected that I misspelled a phrase or word, we can always use the find and replace bar to fix any errors. We can edit, delete add, basically just alter the words we wrote. Similarly, in our DNA, we have the same function. This function is taken on by a system called CRISPR Cas9.
CRISPR is short for;
Clustered
Regularly
Interspaced
Short
Palindromic
Repeats
CRISPR consist of two main components. The Cas9 protein has the ability to cut through DNA and a guide RNA that can recognize the sequence of the DNA that must be edited. To use CRISPR Cas9, scientists first identify the sequence of the human genome that is causing a health problem. Then they create a specific guide RNA to recognize the specific strands of A, C, G, and T’s in the DNA. The guide RNA is attached to the DNA cutting enzyme Cas9. After that, this complex is introduced to the target cells. It locates the target letter sequences and cuts the DNA. At that point, scientists can then edit the existing genome by modifying, deleting, or adding new sequences. This process efficiently makes CRISPR Cas9 an efficient cut-and-paste tool for DNA editing. In the future, scientists hope to use CRISPR Cas9 to develop critical advances in patient care or even cure lifelong diseases.
A New Form of Gene Editing
Let us get the battle started! Now that you are a mini expert in Genome editing, I think you are ready to understand the complexities of this battle. Sequencing, a strong fighter and an essential would side with CRE, the original ruler in hopes of not making too many changes. Epigenetics and CRISPR however would side with DRE, a new force! CRE, understanding its loss immediately backs down, it knows that epigenetics and CRISPR are too powerful of a force.
Woah crazy story Meera. But what exactly are CRE and DRE?
Excellent question! Down below I have outlined what they are, what they do, and why DRE is replacing CRE!
Cre Recombinase
In essence, CRE recombinase is a tyrosine (a tyrosine is a type of amino acid) recombinase enzyme derived from the P1 bacteriophage (a type of bacteria that infects E-Coli). The enzyme uses a topoisomerase I-like mechanism (a single gate mechanism) to carry out site-specific instructions. Now that you know a bit more about CRE, let us dive into DRE recombinase.
DRE Recombinase
CRE has often been recognized as the forefront tool for SSR genome editing. Scientists believe that having another similar tool in our “genetic toolbox” could be a great help to advancing our learnings. Scientists have developed a new version of CRE called the DRE- ROX system. This system includes an Escherichia coli-inducible expression vector based on the temperature-sensitive pSC101 plasmid, a mammalian expression vector based on the CAGGs promoter, a ROX-lacZ reporter embryonic stem (ES) cell line based on targeting at the Rosa26 locus, the accompanying Rosa26-rox reporter mouse line, and a CAGGs-Dre delete mouse line.
That might have been a ton of gibberish to you. Let me help break it down.
Essentially, the DRE-ROX system is composed of many variants like a temperature-sensitive plasmid. Or a CAGGs promoter. Nevertheless, the DRE-ROX system, is nearly as efficient as the CRE Recombinase, casing it to be a great addition to our genetic toolbox. DRE is a newer tool that can help expand scientist's research in many ways and fields. Due to this reason, CRE stepped down from its throne and the DRE-ROX system took over.
Hello! My name is Meera Singhal, and I am a 13-year-old currently fascinated by the field of biotechnology, specifically stem cells and gene editing. I’ve written articles about biotechnology, mindset tips, and a whole variety of up-and-coming topics. Interested? Check out my medium, LinkedIn, YouTube, or TKS Life Portfolio for more content! Curious to see more about me? Consider subscribing to my Newsletter! Thank you so much!
