Molecular Biology is the Transcription from DNA to RNA

(PCR), which isolates small fragments of DNA that have a high degree of variability from

PCR permits the synthesis of millions of copies of a specific nucleotide sequence in a few hours. It can amplify the sequence, even when the targeted sequence makes up less than one part in a million of the total initial sample. Steps of the PCR cycle are shown in below figure.

Since the temperature optimum for the DNA polymerase to act is 72° C, the reaction is heated to that temperature. DNA polymerase lengthen the primers by attaching more nucleotides onto the primer in a sequential manner, using the target DNA as a template.

The primary purpose is to identify a genetic marker or study the function of a specific gene. There are three steps involved in this process which are as follows: denaturation, annealing and elongation. Denaturation involves heating the DNA to agitate the hydrogen bonds, and annealing allows the temperature to be lowered so that the primers can be “annealed” to the single-stranded DNA template. The last step requires DNA polymerase to synthesize a new strand of DNA that is complementary to the RNA strand in the 5’ to 3’ direction (Amplifying DNA: The Polymerase Chain Reaction, 2016). The forward and reverse primers are needed to start the replication process by providing the appropriate nucleotides to the new strand. On the contrary, sanger sequencing makes copies of a target DNA, and the the DNA strand that will be sequenced is separated into two strands, so they can be copied through chemically altered bases. The altered bases cause the process of copying to terminate each time a particular letter is added to the growing DNA chain, which happens to all four bases until the fragments are put together to reveal the original sequence of the original DNA. The aforementioned processes are thoroughly explained to give an overview of the steps involved in providing the end products of the experiment, so an individual can manually decipher

PCR works by denaturing the double-stranded DNA and annealing the primers to the newly-made single-stranded DNA, leading to the extension/elongation of the DNA by a polymerase that attaches to the primer/DNA strand. The PCR reaction strums through a handful of temperature cycles to maximize each step and the amount of product.

A type of next-generation sequencing is illumina sequencing. Illumina sequencing sequences large amounts of DNA in a single attempt. In this process, the sample is firstly cleaved into short fragments. Then, PCR is performed in order for amplification of each read to be carried out. This would subsequently lead to the creation of numerous copies of the same read, at a particular location. Then, separation into single strands occurs, so that sequencing can be performed. This process is followed by the introduction of fluorescently labeled nucleotides and DNA polymerase into the slide, with a termination so that one base is added each time. Therefore, at each read location, a fluorescent signal would be present to indicate the addition of a base. Then, another cycle is performed, with the terminators and fluorescent signals being removed so that another base can be added and to prevent the signal from interfering with future cycles. Computer technology is then utilized to detect the base at each site, so that a sequence can be constructed. This will result in sequence reads of the same length since the same number of cycles will be performed for each

Then, the desired gene is isolated. If the gene comes from a genome that has already been investigated by scientists, there will be a collection of sections of the genome species ready. If the genome where the gene is taken from is unknown, a much more complicated procedure follows, as the collection isn’t there. Most of the time, the genome type has already been used and experimented with. The Polymerase Chain Reaction is used to amplify, or multiply, copies of a DNA piece. It depends on the ability of DNA polymerase to synthesize strands of DNA complementary

A fragment of DNA was then amplified using Polymerase Chain Reaction (PCR) with a forward primer and a reverse primer. PCR is a technique used to make as many copies as possible of a specific section of the DNA extracted. The five necessities for PCR to be successful is the DNA template that one wants to copy, a buffer, nucleotides (ATCG), polymerase, and primers. PCR occurs in 3 stages: (1) Denaturing, (2) Annealing, and (3) Extending. During the Denaturing stage, the stand containing the DNA template (in this case the fetus’) is heated to separate the double stranded DNA. Next during the Annealing stage, the stand is then cooled to allow the primers to attach to a specific location on the single stranded DNA template (primers are usually 20 to 30 bases long). Lastly during the Extending stage, the stand’s temperature is increased to permit the making of new DNA by a specific DNA polymerase. The result is multiple new strands of DNA that has been successfully copied (What is PCR, WEB), followed by electrophoresis in 1% agarose gel. Electrophoresis is a technique commonly used to separate DNA, RNA, and proteins according to their sizes. The result was then purified with a PCR purification kit (removing any remaining DNTPs, or nucleotides that could result in contamination). The products were

PCR is a tool used by those looking to amplify small amounts of DNA for identification purposes. Thermus Aquaticus’ main use is within this DNA amplification process (PCR) is its reproduction enzyme. The bacterium’s polymerase, called taq polymerase (name after the bacterium), is used to reproduce

Transcription is a process in which information in a strand of DNA is copied to a new molecule of messenger RNA. DNA stores genetic material in the nuclei of cells. RNA is a copy of the original DNA material but is not used for long term storage and is free to exit the nucleus. Although it is not an exact copy of the DNA segment. Transcription is carried out by an enzyme called RNA polymerase and proteins called transcription factors. Newly formed mRNA copies of genes serve as blueprints for protein synthesis during translation.

Polymerase chain reaction (PCR) is a technique used in making multiple copies of DNA segment. The director of the Australian Museum mention that Polymerase Chain Reaction (PCR) was a very important step in producing adequate amounts of Tasmanian Tiger DNA in order to proceed the research and complete the cloning project (Research of the Australian Thylacine N.D). Furthermore Geneticists working for the Australian Museum had successfully replicated Thylacine DNA by using this process.

Mullis new method is so significant, he was awarded the Nobel Prize. Polymerase chain reaction is a four step highly specific and rapid method, that can produce billions of copies of DNA, within a test tube in a small amount of time. This process, outlined below, will repeat and as more cycles are completed, the amount of DNA produced will increase as well. The first step in the PCR chain, is the DNA sample itself being heated to the high temperature of 95℃, resulting in the denaturation of the double-stranded DNA, into single-strands. The DNA using two nucleotide primers, is cooled to an approximate temperature of 55℃. Each primer are compatible to each 3’ end of the DNA fragments to magnify it. The reason for the cooling temperature, is because it allows the primers to anneal, with the 3’ end of the single stranded DNA to also be magnified. The third step, in this process, requires the sample to be heated again, but to a slightly less temperature of 72℃, which Mullis found to be the optimal temperature for Taq polymerase, which is a specific kind of DNA polymerase used for the PCR. Without the isolation of the DNA polymerase from the heat-loving bacteria, which ultimately makes PCR able to work. The same as different DNA polymerase enzymes, by adding a nucleotide to the ends of the DNA(using base pairing), it synthesizes. Like stated before, the

The first step, denaturation, uses heat to separate the DNA strand, giving it a single stranded template. Annealing then cools the reaction so that the primers can bind to the complementary sequence of the single strand DNA. During the annealing process, DNA templates can rejoin each other rather than use the primer. To prevent this from happening researchers can decrease the DNA concentration or increase the primer concentration that way the negatively charged DNA template strand can flow towards the positively charged primer. The final step, extension, raises the temperature of the reaction so that the Taq polymerase extends the primer, synthesizing new DNA strands. This cycle continues doe about 20-40 times before the reaction is complete. If the reaction is done correctly, the specific DNA region can be multiplied up to a billion times. This is because the new DNA strand serves as a template for the next set of DNA

Transcription is where DNA is transcribed into RNA which then can be pass to the ribosome’s to act as a template for protein synthesis. Before transcription can begin DNA must unwind and the two halves of the molecule much come apart so exposing the base sequence. This process begins when a region of a two DNA strands is unzipped by enzyme called RNA polymerase attaches to the DNA molecule at the imitation site.

The polymerase chain reaction or PCR for short can be used to create many copies of DNA. This allows the DNA to then be visualized using a dye like ethidium bromide after gel electrophoresis. The process has been refined over the years, however the basic steps are similar.