Lesson Video: Molecular Technologies Biology • Third Year of Secondary School

Video Transcript

In this video, we’ll learn about some different molecular technologies. We’ll discuss bioinformatics and how it can be used to look at evolutionary relationships, so- called gene machines, to produce desired sections of synthetic DNA, as well as DNA hybridization. We’ll discuss the advantages of these technologies and their applications.

We’ll begin by discussing an important molecular technology known as bioinformatics. Humans and chimpanzees share a lot of their DNA in common. In fact, about 99 percent of our DNA is the same. So, how did we determine this? Well, we basically need to line up the two sequences and then look for differences. The problem is, and it’s a big problem, is that the sequence of the genome, or our total DNA and the chimpanzee, is about three billion base pairs long. This is a huge number. It may be easy to write it out on paper, but to give you an idea, one billion seconds is just over 31 years.

So, to make our lives a little easier, we use one of humankind’s greatest tools, the computer. Computers can align these sequences and check for mismatches a lot faster than any human or chimpanzee for that matter is able to. This is an example of the field of bioinformatics. It’s a combination of computer science and biology to analyze large and complex biological data, like our DNA sequence here. So, by using a computer, we can line up the three billion or so base pairs in the genome of the human and chimpanzee, and it will tell us how similar they are in a reasonable amount of time.

Comparing DNA sequences can tell us a lot about how closely or distantly related we are to other species. So, while humans and chimpanzees share about 99 percent of their DNA, what about another species, like a chicken? Now, if we include some of the sequence of the chicken’s DNA, we’ll see that there’s more differences. In the chimpanzee, only one base pair was different from human DNA, which is now underlined in black, whereas in chicken DNA, we can see five base pairs that are different. Because human and chicken DNA is less similar compared to human and chimpanzee DNA, humans and chickens are said to be more distantly related than humans and chimpanzees. This is because humans and chimpanzees share a more recent common ancestor than humans and chickens do and is why humans and chimpanzees have more DNA in common.

Besides looking at DNA sequences, we can also compare the amino acid sequence in proteins to study evolutionary relationships. So, here’s a section of the hormone insulin in our three species, and similar to DNA, bioinformatics can help us figure out the differences in these sequences. Bioinformatics isn’t just about figuring out evolutionary relationships. We can also use it to study gene expression or how proteins fold or how tens of thousands of proteins can interact with each other. There’s a lot going on in a cell that we want to understand, and a computer is the right tool for this job.

Suppose we wanted to study the evolutionary relationship of insulin between these species in more detail. There’s another way we can do this, but before we get into it, we first need to produce a copy of the DNA of the insulin gene. The right tool for this job is a gene machine. A gene machine is a laboratory tool that synthesizes genes or sections of DNA from an input sequence on a computer. You can either put in the DNA sequence directly, or if you only have the amino acid sequence of the protein, then it can work out the DNA sequence from this. Let’s look at this process in more detail.

You’ll recall that DNA is transcribed to make an mRNA sequence, which is then translated to make the amino acid sequence of a protein. So, let’s go through this. Here’s a section of the insulin protein, and if we’re converting this amino acid alanine back into mRNA, we need to look at a codon wheel to do it. So, alanine is located right here, and the corresponding codon for alanine is GCU. The next amino acid is serine, and this corresponds to the codon UCU. Filling in the rest of the codons will give us this mRNA sequence.

Now, we can convert this into the DNA sequence. You’ll recall that mRNA and DNA have directionality, so let’s fill that in. Now, we’ll simply convert this sequence in the DNA, remembering that thiamine in DNA replaces uracil in mRNA. So, there’s one strand. Now, let’s do the complementary second strand. Great! So, there’s our DNA sequence for a segment of the insulin protein. As you’ve seen, we can work this all out by hand, but the gene machine can do this automatically using a computer. Next, the gene machine needs to synthesize this DNA sequence, and to do that, it uses something called an oligonucleotide.

An oligonucleotide is a short piece of synthetic DNA or RNA that is often single stranded. The gene machine assembles two oligonucleotides. The first one corresponds to one part of our sequence on one strand, while the second one corresponds to another part of our target sequence on the opposing strand. You’ll notice that we now have two oligonucleotides that span the length of our target DNA sequence, a portion of the insulin gene. What’s now missing are these two gaps in the sequence at either end. So, how do we fill these in? You’ll notice here we have an overlapping section where the two oligonucleotides meet. This is a region of double-stranded DNA and can act as a binding site for the enzyme Taq polymerase.

Taq polymerase is a type of DNA polymerase, the enzyme that plays a role in replicating DNA in preparation for cell division. We use this enzyme because it’s able to fill in these gaps and can also be used for PCR, or polymerase chain reaction. This is a technique that makes multiple copies of this target sequence, and it’s helpful because we want to have a lot of it so we can extract it and study it. So, Taq polymerase is able to fill in the gap on the top strand by using the opposite strand as a template to add complementary nucleotides, where it can then fall off and then reattach, where it can now fill in the gap on the opposing strand.

We now have our completed DNA sequence synthesized using our gene machine. An obvious advantage of using a gene machine to synthesize genes or sections of DNA is how easy it is. You simply need to input the protein sequence of your favorite protein and it synthesizes the gene for it. Another advantage is that the synthesized DNA is free of introns. You’ll recall that genes often contain introns, which are sections of noncoding DNA that need to be spliced out from the mRNA in order to assemble the protein coding sequence. Since we’re inputting the protein sequence directly into the gene machine, we don’t need to worry about noncoding sequences.

Now that our insulin DNA has been synthesized, we’re ready to study more about the evolutionary relationship between our human, chimpanzee, and chicken insulin. To do this, we’ll look at a molecular technology called hybridization. Hybridization is the process of combining complementary single-stranded DNA or RNA molecules. The two DNA or RNA molecules do not need to be from the same source. So, we could hybridize our human insulin DNA with our chicken DNA to see how similar they are to each other. Let’s go over how this works. So, here is our human insulin DNA segment, and in between the two strands are hydrogen bonds that hold the two strands together.

By heating our DNA typically to 95 or 100 degrees Celsius, we can weaken and ultimately break these bonds, which causes the two strands to separate into single-stranded DNA. And now, if we lower the temperature rapidly to about five to 10 degrees Celsius, the complementary bases will pair with each other again to reform the hydrogen bonds. This pairing of complementary single-stranded DNA or RNA molecules is called annealing. What’s interesting is that these two sources of DNA or RNA don’t need to be the same. So, if we had the corresponding DNA for the chimpanzee insulin gene, we could hybridize these two DNA molecules together. So, we mix these two DNA molecules together then heat the sample to break the hydrogen bond and form single-stranded DNA. And now, we’re going to cool the sample so the single-stranded DNA molecules can anneal.

So, the two single-stranded molecules of human DNA and chimpanzee DNA can come back together and form their original DNA molecule. But sometimes, one of the strands of chimpanzee DNA will anneal to the complementary strand of human DNA. So, let’s see what the hybridization of these two strands looks like. So, here in this human- chimpanzee hybrid of a section of the insulin gene, we can see that most of these bases are complementary to each other and form hydrogen bonds except for one. This one base is different in the chimpanzee DNA, and so this won’t hydrogen-bond. We can see a more dramatic demonstration of this if we hybridize the human DNA to the more distantly related chicken DNA.

Here we can see that there’s many more mismatches. Because there’s fewer hydrogen bonds holding the human and chicken hybrid together, it will take less energy or a lower temperature to separate the two strands compared to the human-chimpanzee hybrid, which has more hydrogen bonding and will therefore require more energy or a higher temperature to separate the two strands. In this way, we can use hybridization to show how similar two sequences of DNA are to each other, which can be used to establish evolutionary relationships between species.

Besides using hybridization to study evolution, it has other applications as well, one of which is called a microarray chip. So, suppose you work with blood donation and thousands of people have donated their blood. Your job is to screen these samples for infectious diseases like HIV or hepatitis. Handling this task one by one would take a very long time. A microarray chip is a little chip that could fit in the palm of your hand and contains thousands of tiny wells that are like little test tubes for hybridization experiments.

Let’s take a closer look at one now. Attached to the bottom of each well are oligonucleotides with a specific sequence. Since we’re talking about infectious diseases, these can be specific to hepatitis, HIV, or any kind of pathogen that causes that disease. So, our blood sample is processed to extract all the DNA or RNA present in the donor blood, which potentially includes the DNA or RNA of our pathogen that we want to detect. This is then labeled with a marker shown here in green.

If the sample contained a pathogen that we’re screening for, then its DNA or RNA will anneal to the complementary oligonucleotide attached to the chip. This will fluoresce and cause the corresponding well in the microarray to light up. We can see the results over here, with each dot representing a different donor, with positive samples being indicated by a green dot. These donors would not be suitable for blood donation.

Now, let’s look at a practice question and apply what we’ve learned in this video.

DNA from different sources can be combined, or hybridized, in a series of steps. Firstly, the double-stranded DNA is broken into single strands. After that, how are the single strands of DNA from different organisms annealed to each other? (A) The enzyme DNA ligase is used to catalyze the formation of peptide bonds. (B) The enzyme DNase is used to repair the broken covalent bonds between bases. (C) The temperature is rapidly increased to provide the energy required for hydrogen bonds between bases to form. (D) The strands are physically forced together until they bind. Or (E) the temperature is cooled so hydrogen bonds between complementary bases can form.

This question is asking us about the steps involved in DNA hybridization. Let’s clear these answer choices so we can have more room to work with. DNA hybridization is the process of combining two complementary single-stranded DNA or RNA molecules and has many useful applications, for instance, in determining evolutionary relationships between organisms.

Suppose a new species of ape was discovered, we’ll call it big foot because of its impressively large feet, and we want to see how closely related we are. We happen to have a bit of their DNA, a portion of which is shown here, and we want to compare the two sequences. One way we can do this is with DNA hybridization. You’ll notice that both strands are indicated, one being the five prime the three prime strand and the other being the three prime the five prime strand. You’ll also notice that these two strands are being held together by hydrogen bonds shown here as these black lines. These hydrogen bonds are holding the two strands together with a certain amount of energy.

By increasing the temperature, we can break these bonds. This then allows a double-stranded DNA molecule to separate into two single-stranded DNA molecules. This process can be reversed by lowering the temperature. This allows the hydrogen bonds to reform between the two strands, and they can anneal or come back together to form the double-stranded molecule. In this case, both of the original strands came back together in the human and big foot DNA. This does happen, but sometimes the big foot DNA will come together with the human DNA too. Because the two sequences aren’t identical, not all of the bases will pair.

Now, if we were to isolate this human-big foot hybrid and increased temperature, because of these mismatches, the energy or temperature required to break these strands apart can be used to estimate how similar the sequences are, which can be used to describe an evolutionary relationship. During DNA hybridization, no enzymes are used, only changes in temperature. And by lowering the temperature, the two single strands of DNA can anneal to each other to form hydrogen bonds.

Now, let’s go over some of the key points that we covered in this video. There’re many different molecular technologies that are commonly used. Bioinformatics uses computers to evaluate biological data like DNA sequences and can be used to help establish evolutionary relationships. Gene machines synthesize genes, or segments of DNA, from protein sequences. And not only does this make obtaining sequences of DNA easy, but these sequences are free of noncoding regions. DNA hybridization can be used to anneal DNA or RNA from two different sources. And this can be used to help establish evolutionary relationships between species or to perform specialized techniques such as a DNA microarray.