GMOs Pt 2: How to Make Your Very Own!

Disclaimer: Trading Atoms has no interests, financial or otherwise, in any biotechnology or related company.

In Part 1 of this series we delved into the realm of genetics and looked at just what constitutes a genetically modified organism (GMO). We said the essential difference is that a GMO usually produces one or more extra proteins that don’t exist in the original species. These extra proteins were added to create some kind of desired trait, such as pesticide resistance in wheat. In the coming instalments we’ll look at the prevalence of GMOs and whether we should be terrified, but before all that dry reality, it’s time to make our very own!

https://tradingatoms.files.wordpress.com/2014/10/166d6-umbukulizard.jpg

The fabled Umbuku lizard.

 

Step 1: Pick your species and desired trait

Back in the idealistic days of my childhood, I had a vision for what my life’s work would be: I would be the one to engineer the world’s very first actual Pokemon! It would probably look something like this.

However, as the years rolled by I gradually came to accept the harsh truth: I would never achieve my dream. The problem was that Pokemon tended to violate the laws of physics. And that was before even considering the technical limitations to genetic engineering. So with this lesson of genetic hubris in mind, what kinds of creatures could we build?

Until quite recently, the limited tools at our disposal for manipulating DNA meant that the best we could aim for was the addition or subtraction of maybe a few genes.

This is no longer the case. With the creation of the first self-replicating synthetic bacterial cell, and the development of new, extremely versatile genetic tools, DNA can be snipped and chopped and changed in pretty much any way we want. Luckily this hasn’t yet lead to an influx of dystopian creations – say, a weaponised psychedelic wasp, or my personal favourite in GM scaremongering:

But if the sky’s the limit when it comes to DNA manipulation, why haven’t we seen this kind of stuff? Let’s assume that scientists are bound by absolutely no ethical qualms or regulatory oversights, and would be keenly interested in adding a digestive tract and muscular system to the common carrot.

The reason, it turns out, is that the way embryos develop is really, really complicated. To make something as complex as a limb, thousands of different genes have to be turned on and off in precisely the right moments in the right cellular locations and at the right levels. Embryonic development is a splendidly complex genetic symphony. Just look how confusing and boring the development of a fruit fly is!

 

 

And that’s a highly simplified explanation, only looking at the very first cell. As you can imagine, the process gets exponentially more complex as different types of cells and tissues begin developing and talking to one another. It quickly reaches the point where a detailed understanding is nearly impossible. The complete story though, if we do ever one day manage to unravel it, looks to be quite beautiful:

So, now that our wildest dreams have been crushed for the foreseeable future, what are we left with?

Well we can still do a lot of pretty interesting things, provided it only involves fiddling with simple and well-understood systems. Generally speaking, this means we’re still limited to changing one or a few genes at a time. While no one is going to be adding wings to lions any time soon, some noteworthy innovations have still been made.

  • One of the earliest breakthroughs, taking place as early as 1978 and providing a major boon for type 1 diabetics, was the insertion of the human insulin gene into E. coli. Before this time, insulin could only be harvested from the pancreatic glands of slaughtered pigs and cattle – not a cheap or pleasant process for anybody. These days, E. coli bacteria happily grow away in vats churning out the stuff.
  • With climate change increasingly impacting upon the yield and yearly predictability of agricultural harvests, drought-resistant wheat may soon prove an important tool in the fight for food security, not to mention farmers’ livelihoods. The wheat is being developed right now.
  • The first genetically modified animal proposed for human consumption is the AquAdvantage salmon. It possesses an extra growth hormone gene that came from a related species of salmon. This extra growth hormone causes it to reach full size in about half the time of a conventional salmon.

 

While all these developments are clearly useful and quite interesting, none of them are very visually exciting. So, without any further delay, let’s see if we can make a cat that glows in the dark. If all goes to plan, here’s what our GlowKitty might look like:

 

 

 

Step 2: Figure out how to obtain your trait

Fiddling with an entire biochemical pathway is Hard, but luckily for us, the modification needed to make a GlowKitty is actually quite simple – we only have to add a single gene. This gene will make a protein called Green Fluorescent Protein (GFP), which looks a bit like a microscopic barrel. The barrel works by absorbing high-energy blue light and re-releasing it as green light. As long as the gene is turned on in enough of the kitty’s cells, we should get a good healthy glow. Note: the gene that makes GFP is also named GFP. This can be a little confusing, but it’s standard practice in the world of genetics.

But where do we get this handy gene from? GFP originally comes from a handsome bioluminescent jellyfish which lives off the coast of North-Western America. Its name is Aequorea victoria, the Crystal Jelly.

As an aside, GFP has probably been played with more than any other gene in history. If you hadn’t heard of it before, you can find it cited in thousands upon thousands of papers. The extremely handy thing about GFP is that you can stick it onto another protein that you’re interested in. Usually, trying to look at a protein in a cell is like trying to spot a black plane in the night sky. Adding GFP is like installing a navigation light.

So, back to the project at hand. We’ve picked our species (cat), decided what trait we want to give it (glow in the dark), and we know we can get the trait by adding a single jellyfish gene (GFP). Time to move on to…

 

 

Step 3: Clone the gene

When Hollywood does genetics, it likes to delve into the spicy issue of cloning things. Things like dinosaurspeople, maybe alien-people (Caution: there may be a spoiler or two in there for anyone living under a particularly stable rock).

There’s also a smaller-scale, less sexy type of cloning you can do: simply copying a piece of DNA. It still counts as cloning! You’re replicating a biological sample aren’t you? It turns out that this kind of cloning is way easier than creating a whole living creature. In fact, it’s an extremely common and straightforward lab procedure, and cloning GFP will be our next step in making GlowKitty.

The process used is called Polymerase Chain Reaction (PCR). If you’re not familiar with PCR, it’s a bit too detailed to explain properly here. Basically though, it involves mixing DNA with enzymes and repeatedly heating and cooling the mixture to help the enzymes copy the DNA. This video provides a pretty good insight into what goes down in the lab whenever somebody does PCR:

 

 

PCR is an amazingly versatile technique. As the song scientific video explains, it’s central to a whole bunch of DNA-related techniques, from paternity testing to detecting mutations and forensic investigations.

Now we’ve covered the theory, you should get out your PCR machine, turn it on and have it idling at about 90-100ºC. If you don’t have a PCR machine, you can substitute in an oven, a bowl of ice and a pair of tongs. Then just follow these easy steps:

  1. Prepare some DNA containing the jellyfish GFP gene.
  2. Add a dash of DNA-copying enzymes (known as “polymerases”). These can be harvested from bacteria, or really any living creature. Make sure to use only trusted species, as cheaper options can result in mutations. Pyrococcus furiosus makes a product that you can count on for peace of mind.
  3. Season with loose DNA bases, salts and primers.
  4. Cook for about two hours, cycling between hot and cool.

Et voila! If all has gone to plan, you should now have several billion copies of your GFP gene.

 

Step 4: Put the gene into your species

We hit an immediately problem here: we can’t just inject the GFP gene into an adult cat. If we did so it would only end up in a few cells, and we want our cat to be glowing all over. We’d also like it to one day be able to have GlowKitties of its own, so we need the gene to be in its sex cells too.

The only option we have is to get the GFP gene into a single-celled embryo. This way, the GFP will join the rest of the cat’s DNA, and when the embryo grows and divides, the GFP gene will get copied into every cell too. So, go out and get your hands on some cat embryos.

There are a range of approaches we could try in order to get our gene in there. Injecting it into embryos with a tiny needle is pretty tedious and finicky, but it does seem to work quite well for a lot of species. We could also try chemicals. There are compounds that punch holes in the outer “skin” of cells, allowing our gene to slip in. The problem is that, unsurprisingly, this tends to seriously weaken the embryos. There are other types of chemicals that wrap DNA up in a ball of fat, allowing it to slip right through the embryo’s skin like a ghost through walls. Unfortunately, these chemicals also tend to be a bit toxic.

Or, there is this:

Yes, that is literally a gene gun. Or if you like, “biolistic particle delivery system”. It fires tiny balls of some kind of heavy metal, often tungsten or gold, which are coated with DNA, right into cells. It works pretty well for plants and animal tissues, where there are a bunch cells together to take the impact. However, as you can imagine, blasting a defenceless little cat embryo with balls of tungsten is like cannonballing a ship. Not good.

As with most things, evolution itself has devised a more elegant solution than anything us humans have been able to come up with. Viruses and certain bacteria have spent billions of years mastering the art of slipping inside living cells. Luckily for us, it’s not too hard to harness these clever critters to do our bidding. We simply have to take out their genetic material and replace it with the GFP gene, and we’ve made the perfect little Trojan horse.

Whichever technique you end up choosing, hopefully you’re successful and get the gene in there.

 

Step 5: The agonising screening process

After all this rigmarole, we might still only be halfway to having our GlowKitty! It’s time to carry out a bunch of screens and checks, not to mention then raising our embryo to an adult cat.

Most life forms have state-of-the-art defence systems to stop new genes from sneaking into their DNA – after all, that’s the kind of nefarious thing that a virus might try to pull. These defences can also make it quite hard for us to get DNA to stay in an embryo. Depending on what technique was used, we might have to screen hundreds or even thousands of embryos to find one that has taken up the gene. This process can be pretty exhausting, especially in something as complex as a cat. Sooner or later though, we should have our glorious eureka moment.

The GFP gene will have picked a spot somewhere along the cat’s DNA to bury in and join the family. Again depending on what technique was used, the spot was probably picked completely randomly. If we’re lucky, it will have picked a boring patch of DNA that wasn’t doing anything. If we’re unlucky though, it might have dived right into one of the cat’s genes and messed it up. There’s also the chance that two or three copies of GFP have jumped in, all at different spots. We definitely need to investigate this, and we do so by reading the DNA code on either side of the GFP gene.

We can compare these DNA sequences to the cat genome to see where the GFP has buried in. If there are no cat genes in these areas, we can be happy that the GFP hasn’t screwed up anything and push ahead. Otherwise, it’s back to the screening process to find a different glowing embryo.

As the cat develops, we’ll have to monitor that the gene is making enough GFP – but not too much – that it’s making it in the right tissues, and that nothing else unexpected has gone wrong. With a bit of luck though, the cat will grow to term happy and healthy and glowing green.

If you’ve made it up to here: Congratulations. You have obtained your GlowKitty.

 

https://i1.wp.com/i.kinja-img.com/gawker-media/image/upload/s--bKtNQ7cq--/18lq421ptf7j0jpg.jpg

scienceinseconds.com

The Twist

If you had your finger on the pulse back in 2011, you may know that GlowKitty already exists!

The story is that U.S. researchers wanted to study the cat version of HIV (called “Feline Immunodeficiency Virus”, FIV). They did this by adding a resistance gene again FIV, and joined it to GFP to act as a beacon. They followed the same process that we have, choosing a virus to get the genes into cat embryos. These cats can now glow in the dark, and won’t get AIDS as easily.

If you’re the type to be upset by this kind of manipulation of animals, I’ve got some bad news for you: GlowKitty is by no means a unique development. For what it’s worth though, glowing in the dark is not thought to cause any pain or emotional distress, and GlowKitties can lead essentially normal lives, probably oblivious to their sciencey superpowers.

*               *              *

Stay tuned for GMOs Pt 3: What the Heck is Out There? We’ll be investigating the prevalence and types of modified creatures that have most come to populate the planet.

GMOs Pt 1: Just What is Genetic Modification?

Disclaimer: Trading Atoms has no interests, financial or otherwise, in any biotechnology or related company.

The development of Genetically Modified Organisms (GMOs) is clearly one of the more controversial issues of our time, with a wellspring of strongly held opinions issuing forth, particularly from the political left. With such widespread distrust and uncertainty amongst concerned citizens, the topic is well and truly ripe for some informed discussion. Riper than a GM Flavr Savr tomato, some might even suggest.

In this first instalment on GMOs, we’ll be going through the basics of just what genetic modification means. Stay tuned for Part 2 where we’ll walk you through a simple guide on how to make your very own GMO, then in Part 3 we’ll address the more sobering question of whether the technology is even safe, and possibly have you regretting that spider-shark you’ve unleashed upon the world.

A Quick Review of Genetics

As you would surely have heard at some time, all living creatures have DNA (deoxyribonucleic acid) in them. If you’d like to get a bit spiritual-sciencey (as we sometimes do), you can legitimately think of DNA as the mystical life force that vibrates through and connects all living creatures on the planet. It is the real-world midichlorians. This particular molecule is present in every single life form, from the elegantly simple bacteria, to the towering trees, to the most majestic of animals.

Capture

Pretty much everything you need to remember about DNA is contained in the following three sentences. DNA is an incredibly long spiral ladder, with four types of rung. These rungs are organised into genes. Each gene is a blueprint to make a certain protein.

When the word ‘protein’ gets mentioned, most people think of that new diet they’re trying, or how sigh, they really should be making better use of that gym membership. While it’s true that muscles are largely made up of two particular types of proteins, there are many, many more types. It’s actually best to think of proteins as tiny machines that swim around in your cells, controlling every single thing you ever do. They are like the little cogs whirring away driving the immense living robot that is your body.

So to recap:

DNA –> is organised into –> Genes –> are blueprints for –> Proteins –> are tiny machines that control everything you do

A protein-machine grabbing onto pink DNA

A protein-machine grabbing onto pink DNA

How Many Genes are There?

Humans are intricately complex beings, with a huge array of different cell types and processes going on. Before the Human Genome Project, scientists speculated about how many different types of genes and proteins we must have to sustain all this complexity. Guesses ranged from over 6 million genes back in the ’60s, down to 100,000 genes by the National Institute of Health in 1990, to a post-genome estimate of 22,000. Recent evidence suggests the number is probably actually around 19,000 to 20,000.

Whatever the exact figure, it’s still very large, especially considering that those sweet guns you’ve been working on are mostly made up of just two proteins. Out of the thousands of others, only a small handful are well understood, and many remain outright mysterious.

How many Genes are there in Other Branches of Life?
kerrydarlington.co.uk

kerrydarlington.co.uk

Bearing in mind that it’s very hard to say exactly how many genes any species has, geneticists have found some interesting results:

Humans – 20,000

Dog/cat – 19,000/20,000

Mouse – 25,000

E. coli4,200

T. vaginalis60,000

HIV – 9    (Note: RNA not DNA)

Brewing yeast – 6,000

Fruit fly – 14,000

Frog – 20,000-21,000

Rice – 46,000-55,000

Wheat – 94,000-96,000

So if you thought that humans were a superior species genetically, think again. While we do have very impressive brains, our gene sets are not so different from a whole bunch of everyday animals. If you’ve ever been unlucky enough to suffer a bout of vaginitis, you may have Trichomonas vaginalis to thank – a single-celled parasite with three times as many genes as you.

Plants in particular can have staggeringly large gene sets. This is often the result of accidental DNA duplications that occur during evolution, which are then chosen by selective breeding (more on that below).

This discussion of gene sets is actually quite facetious, because what has become clear over time is that the number of genes doesn’t really matter. Merely witness the devastation that HIV is able to wreak with its measly nine genes. The important thing to remember is that most species have thousands of genes, and we generally have very little idea what they do.

What Is Genetic Modification?

Time to get serious. Here’s the legal definition:

“Any living organism that possesses a novel combination of genetic material obtained through the use of modern biotechnology.”

This is basically saying that an organism is considered to be a GMO if it has had its DNA changed by scientists. There are a few possible kinds of changes:

  • inserting a gene from another species
  • editing (mutating) an existing gene
  • deleting an existing gene
  • changing how much protein a particular gene makes (regulatory changes)

Overwhelmingly, the GMOs created to date have had one or a few genes added to them from other species to create new functionality. A few examples of this include:

Contrary to the raft of hysterical images circulating online about GM food in particular, GMOs are NOT injected with mysterious chemicals, they do not gain explosive or radioactive properties, and they do not spontaneously develop circulatory systems.

Rather:

GMOs have a tiny difference in the proteins they make

To illustrate this point, let’s say that scientists make a drought-resistant strain of wheat by adding two genes.

The gene sets of the two strains would look like this:

  • Original wheat: 95,000 genes, making 95,000 proteins
  • GM wheat: 95,002 genes, making 95,002 proteins

Your body doesn’t know what any of the original 95,000 proteins are, and we’re not specially adapted to be able to deal with them. Rather, imagine a conveyor belt manned by thousands of eager unsupervised 5-year olds, with intricate Lego creations travelling along it. It’s going to be an orgy of joyful destruction.

Our digestive systems are much like this. Whatever shape or function a protein has, this becomes irrelevant once it enters the stomach. Gastric juices and enzymes will tear apart everything. The two extra wheat proteins will be broken down just like all the others.

It is possible that, while still in the wheat, the drought resistance proteins could make a chemical that is relevant to human health, such as bacteria that produce insulin. For this reason not all GMOs are equal, and the functions of introduced proteins have to be well understood. In most cases though, the only difference between GMOs and “wild” strains will be one or two extra proteins. We’ll explore health risks of GMOs further in Part 3.

What isn’t Genetic Modification?

An organism can be considered GM if even a single rung in its DNA ladder is changed – even if that rung does absolutely nothing. Let’s return to our legal definition:

“Any living organism that possesses a novel combination of genetic material obtained through the use of modern biotechnology.”

The important clause is “through the use of modern biotechnology.” What this means is that the DNA has to be altered in a specific fashion for it to count as genetic modification. Otherwise – bizarrely – any changes are considered natural.

There are several ways that DNA can be altered without the use of modern biotechnology. As we shall see, these “non-GM” methods generally result in far more significant and unpredictable changes.

carrotmuseum.co.uk

carrotmuseum.co.uk

The oldest way that humans have been modifying DNA is through the 10,000 year-old practice of selective breeding. An example of this is cultivating crops with duplicated sets of genes. These plants typically have larger fruit and tens of thousands of newly evolving genes. Humans have also both accidentally and intentionally created hybrid species, throwing together thousands of unfamiliar genes from two species. Modern staple crops, like maize, wheat, rice and fruit trees, are all human-created mutants which differ wildly from their natural ancestors.

A far more rapid process is that of random mutagenesis. If adding one gene using biotechnology was like carefully painting a single dot on a piece of canvas, random mutagenesis is Jackson Pollock. It involves splattering random and sometimes catastrophic changes all throughout a species’ DNA, potentially affecting hundreds of genes at once. This can be achieved chemically with a substance like EMS, but another method frequently used by farmers, “radiation breeding”, simply involves shining a little X-ray or gamma radiation on seeds before planting them. China has even sent seeds to space to give them a nice gamma ray bath.

Predictably enough, random mutagenesis is massively destructive to most of the seeds exposed. However, sometimes a few will mutate in just the right way to gain new functionality such as faster growth or better yield, and these are what farmers are after.

Unlike GM strains created with modern biotechnology – which have to be extensively characterised and regulated – randomly mutagenised seeds are rarely (if ever) characterised, let alone disclosed to consumers as being mutants. Almost no country except Canada has any regulatory restrictions or requirements around the practice, nor does random mutagenesis violate any country’s organic standards.

Thinking about this for a second, we reach an absurd yet true conclusion. It’s completely possible that:

a) A specific mutation could be created in a lab using modern biotechnology. Meanwhile at a farm, completely by chance, the same mutation could be created using random mutagenesis. The resulting two organisms would be identical, but only one of them would ever be characterised, labelled or regulated.
b) An organic company that was fervently against “GMOs” could employ random mutagenesis in their crops. In fact, have you ever bought an organic Rio red grapefruit?

The practice of radiation breeding is on the rise, and possibly far more prevalent than anyone realises. Furthermore, there are solid arguments that conventional GMOs pose less threat than randomly mutagenised seeds. As a result, the current regulatory situation is, to put it politely, extremely strange.

The development of GMOs is an important issue for us to collectively address as we move into the future. Until the science is understood by people like you and me, no informed policy decisions can be made, and we’ll be stuck with the kinds of illogical regulations that currently exist. So if you’ve made it this far, congratulations! You’re a part of the solution, and next time you hear the term “GMO” you can think, “Aha. That means a protein has probably been added.”

Interested for more? Read on with GMOs Pt 2: How to Make Your Very Own!