[Music] hello and welcome to theme D in the IB Biology curriculum continuity and change in this video we are focused at the molecular level and we'll be reviewing the process of DNA replication now if you were asked to replicate something like a painting you would want to gather all of the proper materials for you to make an ident identical copy of the painting down to the Last Detail cells have the ability to do just this but with the long molecule found in the nucleus called DNA the process of DNA replication describes how cells go about taking one strand of DNA and creating an identical copy of it resulting in two strands before we dive into the details of how this works let's first consider the why why would a cell need to replicate its DNA there are two main reasons which are organism growth and reproduction there was a point in time when all of us were just one cell large with one set of DNA packed into our chromosomes but where did we even get this DNA from the answer is our parents we are a mixture of our parents' DNA as half of the chromosomes we get from our dad and the other half from our mom but they didn't just hand over the only DNA they had they made copies of their DNA through replication which ended up in the sperm and egg cells that created us but look in the mirror now and you can easily deduce that you are no longer only one cell large but made up of trillions of cells through the process of growing from one cell to trillions each step of cell division was supported by DNA replication which ensures that all of our cells each have a copy of our original DNA that was given to us by our parents again another process that would not happen without DNA replication and a whole lot of it the process of DNA replication is said to be a semiconservative process process this means that in the process of creating a new DNA strand from an old existing strand the old Strand and New Strand take up an equal percentage of each new DNA molecule when the replication process is done which is 50% original Strand and 50% newly synthesized strand this happens because when the DNA is being copied it needs a template to read and without a template the complimentary bases would not be put together in the correct order to make an exact copy so the old strand gets split in half and becomes a template where nucleotides with a high degree of accuracy attach to fill in the missing pieces the base pairing rules for DNA are always consistent in that adenine pairs with thyine and cytosine pairs with guanine which helps ensure that an accurate copy is created so by the end we conserved half of the original strand in each DNA copy and built up half of each strand with new nucleotides which is why they call it semiconservative so we know a bit about replication like what it does and how the original DNA strand is conserved let's now dive into a few more details about how the process works at the molecular level in order for the process of DNA replication to begin the two strands of the original DNA molecule must be separated this means that the hydrogen bonds that hold the bases together must be broken this separation is performed by the enzyme helicase it moves across the Strand separating the bases by breaking those hydrogen bonds and unwinds the DNA now with the bases on each strand freely exposed another enzyme called DNA polymerase begins to add complimentary bases to the open strands which are acting as a template for the New Strand being built for every adenine base on the old strand the DNA polymerase pulls in a thyine to pair with it and it does the same for guanine and cytosine as they always pair together again the rule of complimentary bases is followed here to ensure that the two strands of DNA end up being identical the DNA helicase will continue to break and unzip the Strand at different locations while many DNA polymerase molecules work to build each new complimentary Strand and when the process is finished the cell is left with two complete copies of [Music] DNA for DNA replication is a natural process that occurs within cells that need to divide but humans have of course found a way to recreate it to an extent in the lab that creates new use cases for the process scientists can make copies or artificially replicate DNA strands through a process called polymerase Chain Reaction or PCR for short this process involves a special machine called a thermal cycler specific components like free floating nucleotides primers and an enzyme called Tac polymerase and is completed through three different steps called denaturing analing and extending and to make it all work we must have a DNA sample we can't replicate DNA without first having a DNA section to replicate after all so we have the DNA sample that we want to make copies of perfect let's talk about how each step of the process works the DNA strand primers free floating nucleotides and an enzyme called Tac polymerase are added to the thermal cycler which is the PCR machine during the denaturation phase the sample is heated up to around 90° C which will separate the hydrogen bonds that hold the two backbones of the DNA strand together with the DNA strand separated the bases are now freely exposed next for the analing phase the sample is cooled down to around 55° C which allows the primers to connect to the DNA sample the primer is specifically chosen beforehand to amplify a desired piece of the DNA usually around 100 base pairs in length then for elongation the last phase the sample is heated to 75° C which allows the TAC prase to attach to the primer and and then move along the Strand attaching the free floating nucleotides when this process is over the one original DNA strand now has two copies the machine then goes through another round of heating and cooling turning those two copies into four and then again into 8 and 16 and so on exponentially this is how in just a matter of hours you can go from one copy of a DNA strand to Millions this can be done with DNA from any species and is a common practice used to identify viruses like a Corona virus during the height of the 2019 2020 pandemic many people were getting tested to see if they had coid 19 the best and most reliable type of test was a specific type of PCR test called a quantitative PCR test or qpcr this involveed taking a sample virus from a subject usually through a mouth swab and using reverse transcripts to create a viral DNA sample from the original RNA found within the virus because the SARS Cove 2 virus only contained RNA not DNA the primers used were designed to only connect to DNA that matched with the sequence created from a CO 19 virus RNA sample this process also used free floating nucleotide bases that had fluorescent tags on them so in this scenario the DNA would have only been Amplified into millions of copies if it in fact was from the correct coid virus because of the primer which would have shown a high level of fluorescence with the nucleotides that were used to replicate it meaning you were in fact positive for having the virus aside from identifying virus components PCR has other use cases when paired with other lab Machinery which we will discuss next gel electroforesis is another process that when paired with PCR has many use cases let's talk about what gel electroforesis is and how it works and then dive into how it's commonly used gel electroforesis is a technique used to separate proteins or fragments of DNA according to size samples are placed in a block of gel and an electric current is applied which causes the samples to move through the gel let's remember that the phosphate group in DNA's backbone causes it to be negatively charged overall this negative charge is therefore attracted to the positive electrode side of the electroforesis machine which pulls it through the gel samples of DNA are loaded into the open well at the top and contain copied fragments of different sizes and when the current is turned on they begin to move towards that positive end smaller proteins or DNA strands will move faster through the porest gel with less resistance and therefore will move farther larger molecules on the other hand move slower and travel less distance this results in samples of different sizes traveling at different speeds and ending up at different distances creating bands in the gel as you can see from this picture there are different bands in different spots showing how the DNA fragment sizes differ with longer fragments towards the top and shorter fragments towards the bottom that have moved further through the gel scientists often use a ladder with known DNA lengths to compare new samples putting these two biotechnology Concepts together scientists can use PCR and gel electroforesis to carry out different applications like DNA profiling DNA profiling Compares DNA from multiple sources with the end goal of identifying a person in this process DNA samples are collected Amplified with PCR and then run through a gel to determine the types of DNA fragments present people who are more related with each other will share a similar d a profile noted with multiple matching bands within an electroforesis gel this is commonly used in criminal investigations when trying to match the DNA of a suspect to DNA found at a crime scene or in paternity cases when trying to deduce who the father of a specific child is in the case of the father about half of the Bands should be identical because the child obtain half of their DNA from the father and the other half from the mother this is of course dependent upon the specific markers they choose to amplify understanding how how both PCR and gel electris work is very important for the IB [Music] exam [Music]