A World In Which We Are The Same
Try to imagine if the closest person next to you was the exact same as you. It might be hard to have a person that’s close to you but hopefully, there’s at least one.
Every single part of you two is suddenly the same thing. Your hair, eyes, personality, and everything else. What’s interesting is that all of that relies on one word, genes.
I’m sure you’ve heard of genes before. Your parents might have said “you got that from me” or “you were born with blessed genes”, but not many people know how it actually works.
What are genes
Let’s first start off with the fundamentals, what is a gene?
Well, there are two ways of explaining that. Some would say that it’s a blueprint or an instruction manual for the body. It’s like everyone’s individual fingerprint, except your fingerprint is also apart of your genetics.
The other way is explained by Wikipedias definition:
“A gene is a sequence of nucleotides in DNA or RNA that encodes the synthesis of a gene product.”
The issue there, is about 50% of those words don’t make any sense. At the same time, that is the best way of thinking about it. So rather than going with the basic, and boring definition let’s go with the complex one.
But instead, I’ll explain what all of those complicated words mean as well as other important things to know when diving into genomics.
Before we start, if you want to learn more about gene-editing, I left a couple of terms/concepts that I talked about throughout the article. It’s at the bottom of the article 😀.
A Quick Lesson on The History of Genomics
By understanding the past of genomics, I think we can truly put it into perspective to show how far we have come with technology. I want to talk about one specific event that really started it all.
In 1990, a group of 20 international research centers joined together to start the world’s biggest biological collaboration, the Human Genome Project or HGP.
This project had the goal of determining the base pairs that make up human DNA. Adding on to that, they wanted to identify and map all of the genes in the human genome from both a physical and a functional standpoint.
As I mentioned before, that’s a lot of genes to map.
They started by dividing human chromosomes into DNA segments of an appropriate size, which were then further subdivided into smaller, overlapping DNA fragments that were sequenced.
The Human Genome Project relied upon the physical map of the human genome established earlier, which served as a platform for generating and analyzing the massive amounts of DNA sequence data that emerged from the shotgun phase.
The second phase of the project called the finishing phase involved filling in gaps and resolving DNA sequences in ambiguous areas not obtained during the shotgun phase.
The point is, when we think about what we use now, this method was extremely inefficient. In fact, this project spanned 15 years and took roughly 3 billion dollars of public funding! It’s truly incredible to see how far we’ve come within the field of genomics.
Now I’m sure a lot of those terms also didn’t make sense, but keep reading because we’re about to get into it!
Fun fact: The HGP produced a very high-quality version of the human genome sequence that is freely available in public databases.
What DNA is and what it’s made of
DNA or deoxyribonucleic acid is one of the two types of nucleic acids (we’ll get to the other one, don’t worry). The function of nucleic acids is to store and express any type of genetic information.
A genome is an organism’s complete set of DNA (or RNA in the case of RNA viruses). Regardless of what type of DNA it is, a fully completed genome means you have mapped the entire organism’s genetic content.
Getting more technical, there are four chemical bases in DNA, A(Adenine), C(Cytosine), G(Guanine), and T(Thymine). Together, they make two sets of base pairs.
In any strand of DNA, Adenine and Thymine make a base pair and Guanine and Cytosine make another base pair.
Chargaff’s rule, also known as the complementary base pairing rule says that A and T will always be together just like C and G.
What acts as the backbone of the DNA is a combination of two molecules, one sugar, and one phosphate. To be even more specific the sugar molecule is called deoxyribose.
A base pair + Phosphate molecule + Sugar molecule = A Nucleotide.
These nucleotides are arranged into two long standards which start to form a spiral. It keeps coiling and coiling into a tightly packed structure called a chromosome.
For every cell, we have 46 chromosomes, half from each parent. If we have 46 chromosomes per cell, think about how many we would have in our body. In case you were curious, if we took out all our DNA and lined it up, it could go to the moon and back 6000 times!
Based on the order of all of the base pairs, a person's genome is made. Think about the number of different combinations there are! In fact, all 8 billion people on the planet share 99.9% of our DNA.
There are so many combinations that it only takes 0.1% of the DNA to distinguish the two most different humans.
Even our closest relatives, chimpanzees share 99% of our DNA which is why by making more connections we can trace back the ancestry of chimpanzees and humans to be from the same thing!
The Second Type Of Nucleic Acid, RNA
Remember how I was talking about how DNA was one of two nucleic acids, the second was is RNA or Ribonucleic acid. RNA can be thought of as a messenger from the DNA to proteins.
Similar to DNA there are four bases that makeup RNA. A(Adenine), C(Cytosine), G(Guanine), U(Uracil). So instead of having thymine, uracil replaces it. Adenine and Uracil go together while Cytosine and Guanine still go together.
In DNA, it’s called a base pair because two nucleobases join together. RNA is single-stranded, meaning that there are no base pairs. Which means that we call RNA’s bases nucleobases.
But to really understand it, we need to know what the three main types of RNA are and why they are so important.
- Messenger RNA or mRNA. This is what the DNA uses first. The messenger RNAs purpose is to transcribe genetic information from the DNA to the amino acids, which make up protein. I’ll go into what transcription is later but the overall purpose of the messenger RNA is to copy the DNA information, store it, and then transport it to the amino acids which will make protein. Well, the question might be, why can’t the DNA do it itself and not use the RNA? The reason is that it can’t leave the nucleus because it risks getting damaged.
- Transfer RNA or tRNA. Next up is the transfer RNA. What’s interesting about this RNA is while the other two RNAs are generally single-stranded, tRNAs fold back into itself in a T shape. Ironic isn’t it? The purpose of tRNA is to bring the amino acids together to form the growing peptide chain. Without going too deep, the growing peptide chain is a link of amino acids which will eventually make up protein. So you could say that tRNA helps bring amino acids together to make protein.
- Ribosomal RNA or rRNA. This one is a little less complicated than the others. The purpose of rRNA is to make ribosomes, in fact, most of the ribosome is made out of rRNA. Both rRNA and tRNA are directly involved with protein synthesis. The rRNA combines with special proteins to form ribosomes.
These are the three main types of RNA, but we’re not done talking about them, later in the article we’re going to discuss what RNAs are involved with the gene-editing tool, CRISPR-Cas9.
The Central Dogma
You would think that all of this RNA and DNA stuff has some type of name to it. It does, and it’s called the central dogma.
Based on the image you can visualize how it works. The word transcription means to copy. The mRNA makes a copy fo the DNA that it is trying to do something with.
Then after that, translation is the process in which protein is made based on the information from the mRNA. This is done using the two RNA’s which I talked about earlier.
To really summarize the last two sections up, the central dogma is the process in which we take genetic information from DNA, transfer it to ribosomes using mRNA, and then make protein using tRNA and rRNA.
Sections of DNA are transcribed into the mRNA which brings the genetic information from the nucleus to the ribosome. The ribosome is made of rRNA and protein. The mRNA is read by the ribosome which brings the tRNA in with the appropriate amino acids to build whatever protein the mRNA is designed for.
Prokaryotic vs Eukaryotic
Everywhere we go, there are 2 different types of cells, prokaryotic and eukaryotic. Any cell will be classified as either one. So what’s are they and how are they different from each other?
In eukaryotic cells, most of our DNA can be found in the nucleus of each cell but small bits can be found in the mitochondria which are called mitochondrial DNA or mtDNA.
Anything that is classified as a eukaryote usually is multicellular, meaning that there are multiple different cells. Some eukaryotic organisms include humans, animals, and plants. There are some exceptions where the organism is unicellular.
On the other hand for prokaryotes, all cells are unicellular which means there is only one cell in the organism. Examples include all bacteria and archaea.
But when we look at prokaryotic cells, there is no nucleus or any other organelles so the DNA resides in a place called the nucleoid.
The reason for this is that eukaryotic cells have membrane-bound structures/organelles while prokaryotes have none.
Think of eukaryotes as a full floor in a house and prokaryotes as one room. On the floor, there are a lot of rooms that are separated by walls. This description is also fitting considering eukaryotic cells tend to be bigger than prokaryotic.
Similarly, eukaryotic cells have walls that are called membrane that separates organelles. Whereas prokaryotic cells just have a couple of things in the cell that isn’t blocked by a wall, similar to one room.
This is an extremely important topic in the field of genomics. I would actually say that it is what we want to do in genomics. Gene sequencing is all about figuring out the order of nucleotides in the DNA.
It’s ordered by letters or sometimes colours, depending on the system. A specific colour is associated with a letter. In the example above, blue means the letter is A, C is red, T is yellow and G is green.
Throughout our history, there have been 2 main methods that have been used to sequence genes. The Sanger Method, and the high-throughput sequencing (HTS) techniques.
Both of these methods are used in different situations and it just depends on what the requirements are.
Sanger sequencing is now used mostly for de novo initial sequencing of a DNA molecule to obtain the primary sequence data for an organism or gene. Pretty much, The Sanger Method is often used as a way to validate the HTS techniques.
On the other hand, HTS allows the use of DNA sequencing to understand single-nucleotide polymorphisms — among the most common types of genetic variation within a population.
Polymorphism is the occurrence of several different forms/types of individuals among the members of a single species because of a discontinuous genetic variation.
How we can change genes using gene-editing
Think about it, our genes make us who we are. Physically and mentally it is the instruction manual behind what we can and can’t do.
So that begs the question, what if we could change those genes to do what we want it to do? What if the weakest person on the planet could all of a sudden become the strongest? Or the most unhealthy food becomes the most healthy?
We are barely scratching the surface of what potential gene-editing technology has. Realistically, the possibilities are endless. So doesn’t it make sense that we use this god-like technology to really make a difference in the world? Well, yeah. Later in the article, I’m going to talk about the limitations that hold it back from its full potential.
But before I go into that, I think it would make sense as to why this technology is considered so revolutionary.
Implications of the technology
The scary but impressive part of this technology is it can solve the biggest root causes of some of the world's biggest problems.
The ability to alter a humans gene is something we could use to cure diseases like Cancer, Malaria, and other genetically related diseases. Keep in mind cancer alone is the 2nd biggest cause of death in the entire world.
To be able to solve that with something that is like a copy and paste editor is mind-blowing!
Think of something like world hunger or inefficient agricultural systems. These are some of the biggest issues society is facing. By altering the genes of food to be more nutritious we solve one of the three root causes of world hunger which are affordability, accessibility, and nutrition.
We can make crops require less water which would bring the massive 70% of global water that is used in agriculture down by a significant amount.
CRISPR-Cas9 is a gene-editing tool that is used to mutate genes based on anything you want. CRISPR can be used to cut bad DNA, but it can also be used to add something better.
Think of CRISPR like the control/command F function you use when you’re trying to find what you want in a piece of text. That same function in genomics are technologies like CRISPR-Cas9.
The technology has 2 components. The first component is the Cas9 enzyme that acts as molecular scissors that cut DNA. The second is a guide RNA that recognizes what sequence that needs to be edited.
So you might be asking, how does a guide RNA differ from what we were talking about earlier. Well, the answer is that the 3 RNAs mentioned previously have to do with how the body functions, they are more natural.
On the other hand, guide RNA’s are programmed with a certain purpose and are designed pretty much exclusively for the CRISPR system.
With that out of the way let’s go on to how the CRISPR system actually works.
The first part of CRISPR is to identify where is the issue in the current DNA. If it’s not an issue but rather they want to make something better, they would find where the DNA for that specific thing is stored.
This is where the gene sequencing I was talking about earlier comes in. We have to first map out the genome and then we can say for example “oh the issue is in the following: ATCCGTGCGATTA, now let’s fix it using CRISPR”.
Then, once we identify what letters/colours make up the DNA, we make a guide RNA that is designed to contain the complementary base pair, for every single letter in the target DNA.
In case you don’t remember, a complementary base pair is a base that matches with a corresponding base. So they are C(Cytosine)/G(Guanine) & A(Adenine)/T(Thymine) or U(Uracil) depending on the situation.
Once the guide RNA has made a “copy” of the DNA by using the complementary base pairs, it is attached to the DNA cutting enzyme, CAS-9. Then the guide RNA and the CAS-9 are put into the target cells that we want to fix/change.
Using our guide RNA, we are able to locate the area in the target cell that we want to do something about. This is because we designed the RNA so that it will complement the target area through having complementary base pairs, meaning it can locate it.
After we locate it, the CAS-9 enzyme cuts the DNA in what is called a double-stranded break or the cleavage, meaning it cuts both sides of the DNA. In fact, this is one unique thing about CRISPR and why it’s so hyped up. Double-stranded cutting plays a huge role in CRISPR being more efficient and faster than other methods.
The last part is after we remove the bad gene, it’s just a matter of modifying, deleting, or inserting DNA. This makes CRISPR almost like a copy and paste tool.
In case you were wondering, PAM or Protospacer adjacent motif is a 2–6-base pair DNA sequence immediately following the target DNA. It’s something that really important for the system to have in order to cut the DNA.
This is how we can use CRISPR-Cas9 to edit, remove, or manipulate any gene to our preference.
Note: Sometimes you’ll see the word crRNA which stands for CRISPR RNA which is pretty much the same thing as a guide RNA.
CRISPR-Cas9 is currently the most dominant form of CRISPR and gene editing in general. We’ve gone through a bunch of different methods before it and after but most people consider the Cas9 version of CRISPR to best.
But just like everything else, CRISPR has a couple of issues that come along with it. For example, it can only recognize genetic sequences of around 20 bases long, meaning that longer sequences cannot be targeted.
More significantly, the enzyme still sometimes cuts in the wrong place. Figuring out why this is will be a significant breakthrough in itself — fixing it will be even bigger.
Then there’s also the issue that CRISPR didn’t work well in human embryos and its more recent links to cancer.
Cas12a is an alternative enzyme to Cas9 that is thought of to be a more specific version. Cas12a is also capable of processing its own guide RNA which is something that Cas-9 can’t do. Cas12a also only needs one guide RNA while Cas9 needs 2.
Unfortunately, because Cas12a is not fully developed, the detailed mechanisms of target searching and DNA cleavage by Cas12a are still unclear.
There are a couple of other differences and if you are interested to learn more I recommend checking this out! Just to sum it up, Cas12a is potentially better than Cas9 because it increases the speed and efficiency that CRISPR operates at.
Alternate Gene Editing Tools
Even though CRISPR is the leader in the gene-editing world, it’s still important to know alternative methods that exist.
One of them is called TALEN or Transcription activator-like effector nuclease. This does something similar to what CRISPR does.
It uses a protein called TALE or Transcription activator-like effector to bind to the DNA at a specific nucleotide. Every nucleotide has its own individual TALE(haha so funny). You could think of TALE as a method that is similar to how the guide RNA does that in CRISPR.
It’s also important to note that for every 9 base pairs or so, they join together to make a bigger protein. What this does is it acts as a verification method. This means that the TALE will not bind unless it sees the same, 9 base pair pattern.
After that, we attach half of an endonuclease enzyme called Fok1 to the protein that we made from base pairs(to understand the structure look at the image below). This is similar to CAS-9 because they both cut the desired DNA.
Then we attach the other half to the other side of the DNA and add the same amount of tails but for the other side. This effectively leaves a gap in the middle of the DNA where the Fok1 is.
The last step is the Fok1 cuts the remaining DNA that is in the middle. Keep in mind this is a fairly high-level explanation but does sum up how TALEN works.
Another well known gene-editing tool is ZFN or Zinc Finger Nucleases. Just like both other gene-editing technologies, ZFN uses a binding tool and a cleavage tool.
The DNA binding tool is a protein called Zinc Finger. It can recognize 3 DNA base pairs. In case you didn’t know, 3 DNA base pairs equal one amino acid. If you don’t remember, an amino acid is like a building block that is used to make protein.
When editing the gene, there will be usually 2–3 Zinc Fingers attached to each other which will be able to detect more and more amino acids/base pairs. Having more Zinc Fingers makes it more accurate while reducing off-target recognition.
We then attach a cleavage tool that happens to be the same one that TALEN uses, Fok1, which is a type 2 restriction enzyme. Just like TALEN, we only use half of Fok1 per side.
The reason is that, as a whole, Fok1 is a nonspecific enzyme which means there is no specificity as to where it will cut the DNA, it’s more random.
Obviously, when dealing with DNA, we don’t want that. The goal is for it to be as accurate as possible. When we cut it in half, it becomes inactive. So wait then, how do we get it to cut the DNA when the tool that we’re using to cut it won’t work?
The answer is because we use the other half on the other side of the DNA(to get a visual representation, see the diagram below.)
When these two are aligned they dimerize and cut the DNA. The word dimerize kind of means to merge. By slicing a Fok1 in half, we eliminate the possibility of randomly cutting a gene.
After the gene is gone, the DNA will repair itself which will effectively delete the bad gene and replace it with good, healthy ones.
So why is CRISPR considered better when they all do the same thing?
The best way of doing this would be to compare in a one on one way rather than all three at the same time. I’ll start off with ZFN vs TALEN and then the better one of those two will be compared to CRISPR.
Generally, TALEN is preferred over ZFN despite how similar they are in structure and function.
There are a lot of underlying things that stem off from the main reason. For the most part, people prefer the TALEN method because it can bind to a single nucleotide whereas ZFN binds to three at a time.
This means that it’s much easier to contract a TALEN gene-editing tool compared to ZFN because it’s much easier to change to your liking.
On the other hand, ZFN is limited to its three-nucleotide at a time approach which significantly decreases the changes you can make.
CRISPR vs TALEN
Now that we’ve identified why TALEN is more preferred than ZFN, let’s take a look at CRISPR and its comparisons to TALEN.
Multiple studies have shown that CRISPR is much more cost-effective than TALEN. Most studies show that CRISPR is 3–6 fold cheaper per reaction than TALENs.
But let’s also look at it from a science perspective:
- Simplicity. With CRISPR, the target specificity relies on ribonucleotide complex formation and not protein/DNA recognition. Guide RNAs can be designed in a simple and cheap way to target nearly any sequence in the genome.
- Efficiency. The system is super-efficient. Modifications can be introduced to the DNA by directly injecting the Cas protein and the guide RNA into developing embryos. Usually, it’s a mouse embryo.
- Multiple mutations. Something that’s unique about CRISPR is that genetic mutations can be introduced to multiple genes at the same time by injecting them with multiple guide RNAs. AX Assistant Professor Dr. Haoyi Wang and his team reported using the CRISPR system to successfully introduce mutations to five different genes of a mouse, simultaneously.
These aren't just small pros, they’re incredibly important for the future of gene-editing. When most of us look for a better system in anything, we look for something along these lines.
I’ll talk about this more in the ethics section but all three of these gene-editing methods have one main issue in common, which is that there are off-site effects.
If we can fix this issue with something like CRISPR, we solve the biggest issue with modern gene-editing technology which would bring us a huge step closer to making it a reality.
When we see something that has a new approach and has facts to show that it has so much potential, we jump on it. With CRISPR, it’s no different. But I do think it’s for a good reason.
Ethical Questions and Current Limitations
When we see a technology this incredible, it seems like no matter how bad the limitations are, it wouldn’t matter. To think of the number of possibilities that open when this technology becomes a reality is incredible.
So then why after all these years of this technology being known to do we continue to not use it. Why isn’t CRISPR-Cas9 being used in every hospital to cure the most severe diseases or make superfoods?
I mean there are a lot of answers to a question like that but they all stem from 3 main ones.
One of the biggest ones that have been discussed since the invention of gene-editing technologies is the side effects.
You can change the design plants to operate in fields where less water is necessary for survival, but then need to counter that issue by taking the crops out of direct sunlight because it becomes sensitive to UVA/UVB rays.
It’s the same logic with the human body, when you change a specific gene, whatever genes are related to that will also change, whether that’s good or bad.
In fact, most people would agree that the side effects of gene editing are the biggest reason as to why we shouldn’t use it everywhere we go.
Probably the next biggest concern is that this technology will fall into the wrong hands and would be used for things like wars.
The sad truth is that while we make it infinitely easier to cure cancer or make foods that give incredible nutrition which has the potential to solve some of the biggest problems.
But we can’t only look at the good part. Just as much as gene editing lifts us up, when given to the wrong people, it will bring us down by the same amount if not more.
Just imagine the kind of power you would have if you were able to take anything and change it to whatever you want. Alter the genetic code of anything that has one in such a short amount of time.
Also, even if it didn’t get into the wrong hands, who would have the rights to it? How would we be able to maintain equity and equality throughout the world when certain groups/people have such powerful technology?
I think while gene-editing can be used to solve some of the world's biggest problems, it still has a long way to go in order to deal with the concerns.
Changing the reality of life is unethical
A lot of people think that changing the fundamental parts of our life is not ok because it goes against the practices that we’ve done throughout our history.
This is much more of a perspective issue. To some people, everything should be kept the same as it always has and we shouldn’t tamper with what the fundamental ways of living life have been.
But to others, it doesn’t matter and it’s just about what makes our society better. However, we still need to remember the two reasons above as other justifications for why we aren’t ready to use gene-editing yet.
Important Terms To Remember
This section is going to be kind of like a TL;DR except more focused on learning more rather than summarizing everything. There were a lot of different concepts/terms talked about in the article so this contains the most important ones.
These are in no specific order, ok, let’s get started!
- Fundamentals of Genomics: Genome, DNA, mRNA, tRNA, rRNA, Base Pairs/Bases, Complementary Base Pairs, Phosphate Molecule, Sugar/ Deoxyribose molecule, Nucleic acids, Nucleotides, Chromosomes, Gene Sequencing, Nucleobases, Transcription, Translation, Central Dogma, Ribosome, Prokaryotic, Eukaryotic
- CRISPR Gene-Editing: Cas-9, Cas12a, Cas13, Guide RNA, PAM(Protospacer adjacent motif), Double-Stranded Break, Ribonucleotide
- Miscellaneous: Talen, ZFC, Human Genome Project, High-throughput sequencing, The Sanger Method, Polymorphism, Bacteria Multiplication. Peptide Chain, Cell Membrane, TALE, Zinc Finger, Fok1, Multicellular, Unicellular
I think what’s incredible about gene-editing is that aside from the side effects, all the main issues have to with who gets to use it. I don’t see that as a bad thing necessarily, in fact, it makes me admire how there is no other technology like it.
Think of anything else that is this powerful, to the point where almost every issue is just who gets to use it. These are valid concerns but also does show the immense power that gene editing has.
The current problems of gene-editing are ones that we have to solve in order for this technology to become a reality.
But if we do, we could possibly be looking at the biggest shot we have of winning the war against diseases that humans have been battling for centuries.