Lab 3 – DNA extraction and visualization Journal -Madhu Thalari. 1.Describe the laboratory exercise as you interpreted it.? Ans: This lab has given me methods to extrct DNA from both plant cells and animal cells. The main steps that are followed in both methods made me understand the reasons behind them. In order to extract DNA we need break the barriers(cell wall and cellmembrane), remove water, protiens and other unwanted material, make sure that the chemical we used should not damage DNA that we need and add flouroscent material to visualize the DNA.
n.d.). DNA samples are submitted to a certified laboratory and undergo the following process (DNA Evidence. n.d.): • Extraction is the process of releasing the DNA from the cell. • Quantitation is the process of determining how much DNA you have. • Amplification is the process of producing multiple copies of the DNA in order to characterize it.
In 1948, Linus Pauling discovered that many proteins take the shape of a helix. At Cambridge University, James Watson and his research partner Francis Crick had become interested in Linus Paling’s work. Their approach was to try to make a physical model of what DNA looks like to narrow down the possibilities and eventually create an accurate model of the molecule. In 1951, Watson attended Rosalind Franklin’s lecture on the current work that she had done. Rosalind discovered that DNA could exist in two forms and also discovered that within her x-Ray of DNA, the wet form of DNA had all the characteristics of a helix.
For this reason the DNA is denatured into single strands by incubation with sodium hydroxide solution (NaOH). If the DNA fragments are larger than 15kb, the gel can be treated with acid such as dilute HCl prior to blotting. The DNA is depurinated by the acid and breaks down into smaller pieces which allows more efficient transfer from the gel to the membrane. The Southern Blot Method begins with the DNA fragments being transferred to a nitrocellulose membrane. This is a sheet of special blotting paper.
There are three kinds of DNA with specific functions. First, messenger RNA (mRNA) is responsible for transcribing the information from the DNA. That information is then sent to the ribosomes and cytoplasm. The second, transfer RNA (tRNA) transfers amino acids to the mRNA in a ribosome. The third, ribosomal RNA (rRNA), translates the information from the mRNA and the tRNA.
therefore this process of DNA Profiling can be seen as a useful method of identification with marginal room for error. 1.1. DNA Profiling/Database: The concept is such that, DNA samples are taken from an individual and it is analysed in a laboratory. Further, based on this analysis,
In Fred Sanger Method, the DNA to be sequenced serves as a template for DNA synthesis. A DNA primer is needed which is designed to be a starting point or the initiating point for DNA synthesis on the strand of the DNA to be sequenced. The primer is hydrogen bonded to the 3 ' end of the DNA to be sequenced. The DNA with the primer is divided into four separate reaction mixtures. Each reaction mixture contains all four dNTPs and in addition, one of the four dideoxy analogs (dideoxyribonucleoside triphosphates ddNTPs) of the deoxyribonucleoside triphosphates.
Deoxyribonucleic acid (DNA) is a molecule found in all forms of life that is passed down from parents to offspring. What makes each DNA unique is the chemical makeup of the molecule sometimes referred to as the “blueprint of life.” (BIO). DNA is made up of nucleotides consisting of a sugar, a phosphate and a base pair. About six million nucleotide base pairs make up DNA in each cell. Retrieving this amount of data is both exhausting and time consuming.
After his discovery other method were found and that’s why RFLP is used rarely. How is DNA applied? There are three methods of DNA profiling: RFLP, PCR and STR. RFLP: DNA is cut by restriction enzymes according to repeated sequences of DNA bases, which is unique for every person. The segments taken are separated using a laboratory technique called electrophoresis.
Polymerase Chain Reaction (PCR) is a laboratory technique used in molecular biology to generate a small section of DNA or genes, PCR uses primers to amplify specific genomic DNA sequences with the help of special enzymes, PCR process uses short sequences of DNA and primers and selects specific chromosomes of DNA for replication. (McPherson and Møller, 2009) In addition, there are three main steps involve PCR process, first step of PCR is denaturing when DNA is heated to 90-95 degrees Celsius and double DNA strand is separated in to two single strands DNA, these single strand DNA will act as a template for the new strand DNA, because of the higher temperature the hydrogen bond which holds the complementary strands of DNA together is broken down at this step. (Wheeler et al., 2004) The second stage in the PCR cycle is annealing, this step the temperature is cooled to 50-56 degrees Celsius so the DNA primers can attach to the template DNA by the hydrogen bonding. The final stage of PCR is extending stage when the temperature is raised to 72 degrees Celsius and a new strand of DNA is made by an enzyme called Taq DNA polymerase which is taken from the heat-loving bacteria Thermus aquaticus. This three-thermal process is repeated 20-40 times to produce many copies of the interested DNA sequences.