Definition of DNA amplification Essay

In this experiment, DNA sample from the last week experiment, the DNA extraction is used. The main objective is to amplify the 16S rRNA. It is single print of prokaryote and is very important in the synthesis of protein. DNA is transcript into the RNA by transcription.
The DNA sample is amplified by using the technique of polymerase chain reaction (PCR). Amplify the DNA means make copies of DNA. PCR is done in vitro and it is a method for amplifying a segment of DNA by using the enzyme DNA polymerase. DNA polymerase was purified from Thermus aquaticus from hot spring which the temperature is about 75degreesC to 89degreesC. The highly thermostable DNA polymerase from Thermus aquaticus (Taq) is ideal for both manual and automated DNA sequencing and it is active on wide range of temperatures (Innis, M. A., et al., 1988). The synthesis of DNA is done from deoxynucleotide substrates on a single-stranded DNA template. (Aryal, S., 2015). PCR thermocycler machine is used to do PCR process. It can run 96 samples in 1 run. It consists of gold plate to stand the temperature. The temperature profile is display on the screen of the PCR machine. This process is done in high temperature. For the reaction condition in PCR, the temperature must be correct with each reaction. For example, the temperature for pre-denaturation is 94degreesC, for denaturation is 95degreesC, for annealing 55degreesC, for extension and final extension both is 72degreesC. The cycle is run with 1, 30, 30, 30, 1 cycle respectively for each process. For the pre-denaturation, it is done to adaption for the denaturation. The denaturation reaction causes the heat to break the bond. It causes the double strand broke into single strand. When it is cooled, the denatured DNA will bind again. For the primer anneal, the temperature condition depends on the length of the primer. The longer the primer, higher the temperature needed for annealing. The anneal temperature 45degreesC to 55degreesC is specific to gene 16S rRNA. In the process, PCR buffer, forward and reverse primer is added. The buffer is function to create optimal conditions for activity of Taq DNA polymerase. Primers are required to amplify the target sequences which are oligo-nucleotide with constant length that is shorter than target sequence (Yao, X., 2004). The primer take the bases C, G, T, A from the dNTPs agent. The other technique used in this experiment is electrophoresis. This technique is usually used in the diagnostic applications for infectious diseases, genetic diseases, forensics analysis and so on. It is a method for molecular separation which involved the charged molecules with a direct current (Lee, M., 2009).
the gloves should be wear when handling the reagent. Some of the reagent might be harmful to skin. The EtBr reagent contained carcinogen and can cause mutation, so the gloves must be wear during the handle this reagent. The technique of using a pipette also must be correct and the volume taken also must be accurate so that the best result is obtained.