Practical Techniques in Genomics and Proteomics - Assignment Example

Practical Techniques in Genomics and Proteomics Respond to Question It is importance to chelate metal ions from solution in the process of boiling step at 100 degrees Celsius because chelating agents are chemicals that often form complex, and soluble, molecules with specific metal ions (Ashmead & DeWayne, 1993). They are essential since they help inactivate the ions thus making them not to be able to react normally with other ions and elements in order to produce scale or precipitates. This means failure to use a chelating agent such as instaGene matrix might cause the ions to remain active and, therefore, they are likely to fail to react as expected with other ions. This way, there shall be no precipitates. Response to Question 2 In order to carry out a PCR, one requires DNA. This means a cell is important since it is the source of the DNA. The cell can also serve as a source for base pairs, enzymes and primers. Response to Question 3 There are various structures that are often broken in order to release the DNA from a cell. These include the cells, the cheek tissues, the phosholids, the nucleus, and proteinase K. this is done so that proteins along with other organically-soluble compounds are removed from chromosomes and cells. PCR Amplication. Response to question 1. It is important to have a primer on either side of the segment of a DNA that is going to be amplified because these form the four basic bases. Uracil exists in RNA, despite being a special case. Primers are vital and essential components of the synthesis. They synthesize an RNA short piece which is complementary to the strant of the template DNA forming the bonds of hydrogen with it. This provides a DNA polymer with a starting poing which that is required for the synthesis. After the completion of the DNA synthesis, the segment of RNA is removed and a DNA replaces it. Response to Question 2 Taq DNA polymerase is the thermostable DNA polymerase that derived its name from the thermophilic bacterium called Thermus aquaticus. The T.aquaticus is a specific bacterium found in hydrothermal and hot springs vents. Response to Question 3 It is because they are the only four basic bases with uracil being is present in the RNA, although it is a special case. Components of Master mix include water, oug template, the buffer, deoxynucleotides, aum primers, template DNA and ampliTaq polymerase. The buffer is used for keeping the master mix at a suitable PH in order to make sure the reaction of the PCR occurs. Deoxynu-cleotides, in this case help in providing the energy and the nucleosides used for synthesizing the DNA. As widely cited, it is vital adding equal amounts of each of the nucleotide (, dTTP, dCTP, dATP and dGTP) onto the master matrix for purposes of preventing the mismatch of bases. The ouM primers are short pieces of the DNA that often bind onto the DNA template to allow the Taq DNA polymerase enzyme to start the incorporation of deoxynucleotides. The AmpliTaq polymerase is used, in this case, to add the deoxynucleotides onto the DNA template. Last but not least, there is presence of oug template that is often amplified by a PCR reaction. Response to Question 4 The DNA PCR amplification takes place in repeated cycles of the three temperature-dependent steps. In the first step, a double-stranded DNA or (dsDNA) template becomes denatured. In the second stage, the oligonucleotide primers become annealed to a single-stranded DNA also called (ssDNA) template. In this case one primer become designed specifically to anneal onto a certain specific region found on the left side of the strands of the DNA with the other primer being designed specifically to anneal onto a specific region of the complementary DNA strand right side. In the third stage, the DNA polymerase is meant to extend on each primer in 3 to 5 direction, while duplicating the fragment of the DNA between the primers. While each of the cycle denatures, synthesis and annealing the specific DNA fragment becomes amplified exponentially. Response to Question 5 The precise length of the DNA sequence is not amplied until it is in the third phase because the oligonucleotide primers become annealed to a single-stranded DNA also called (ssDNA) template. In this case one primer become designed specifically to anneal onto a certain specific region found on the left side of the strands of the DNA with the other primer being designed specifically to anneal onto a specific region of the complementary DNA strand right side. Response to Question 1 It is worth noting that Introns and Exons are both related to genes. Whereas the exon is the nucleic acid sequence often represented within the RNA molecule, the Introns, is the nucleotide sequences often found within the genes that are extracted through the splicing of the RNA for purposes of generating a mature molecule of mature the RNA. Introns are commonly found in geneome of higher vertebrae, while introns are less likely to be found in genomes of some verieties of the eukaryotic micro-organisms. Moreover, introns are somewhat less conserved implying that the sequence of introns changes quite frequently with time. Contrally to this, the exons are said to be much conserved implying that their sequence do not seem to change rapidly with time. Response to Question 2 This happens in case the PCR being run is an RT-PCR and the band that has 300 extra bp is caused having a certain contaminating gDNA within the reaction. In this regard, most of the primers for the RT-PCR are such that they are designed to sit in somewhat different exons. This, thus imply that, in case the intron found in between was to be about 300bp lengthwise and the gDNA is then added to the reaction, and to the cDNA then it is certain that two bands could result. These are the shorter or lighter one from cDNA, as well as the longer or heavier band from gDNA. Response to Question 3 There is a reason why the Agarose electrophoresis separates the DNA fragment. This is because the gel has a specific density that is used in pulling the DNA through the gel. A smaller fragment moves faster than the larger one since a spliced DNA is somewhat charged and it starts moving through the gel. Given the density of the gel, it causes that larger piece to move faster compared to the smaller one. References Ashmead, H., & DeWayne, H., 1993. The Roles of Amino Acid Chelates in Animal Nutrition. Westwood: Noyes Publications. Read More