Nucleic Acid Question and Answer

A. DNA Extraction Virtual Lab [2 marks] Question 1 [1.0 mark] Isolate nucleated cells into eppendorf tube (correct starting step). Add 500 ul 10% SDS and 55 ul proteinase K (10 mg/ml stock). Incubate at 37ºC with gentle mixing or rocking. Add 1.4 ml saturated NaCl solution (approximately 6M). Spin eppendorf tubes at 10000 rpm in a centrifuge for 15 minutes. Shake the tube vigorously for 15 seconds to allow protein to precipitate. Transfer the supernatant to another eppendorf tube, leaving behind the precipitated protein pellet Add exactly two volumes of 100% isopropanol at room temperature. Spin eppendorf tubes at 2500 rpm in a centrifuge for 15 minutes. Invert the tube several times until the DNA precipitate is visible. Remove supernatant from tube, leaving behind the precipitated DNA pellet. Dissolve DNA pellet in small volume of TE (Tris-EDTA) buffer or water (correct ending step). Question 2 [1.0 mark] When TE buffer or water is added, the DNA pellet is able to dissolve. However, in the presence of ethanol, the DNA will precipitate. This is because, unlike water, ethanol has a low dielectric constant as it is less polar. That means that Na+ and PO3- could interact with each other more easily and cause the DNA to become less hydrophilic and be precipitated instead. B. Gel Electrophoresis Virtual Lab [2 marks] Question 1 [1.0 mark]

The pieces of DNA have to move through the gel, where there will be some resistance. Large DNA fragments will face more difficulty in doing so as they cannot slip through the holes easily. Higher concentration of the gel will cause the hole size to decrease so the DNA cannot pass through it easily. It will be slowed down and travel an even shorter distance or not be able to travel much at all. This could lead to inaccurate or unreadable results. Question 2 [1.0 mark] Dye is used so as to better observe and track the movement of the particles during electrophoresis. This is because we cannot see the DNA colour with naked eye. The loading dye will help weigh down the DNA so it will sink the bottom of the gel and not drift. C. PCR Virtual Lab [2 marks] Question 1 [1.0 mark] Extracted DNA: this is important so that we can replicate the DNA more times. It is used as a template. Primer 1 : Primer attach to the sites on the DNA strands that will be amplified so that they can copy specific DNA sequences without targeting the wrong site. So Primer 1 will attach to the first site (the start).

Primer 2: Primer 2 will attach to the second site (the end). Nucleotides: Forms the base that makes up the DNA code. DNA polymerase: Attaches the DNA code it reads to the matching nucleotide to make multiple copies of the DNA. Question 2 [1.0 mark] EDTA acts as a chelating agent. It binds cations and prevents enzymes from binding to the DNA. With increased concentration of the EDTA, there may not be good reaction conditions for DNA polymerases due to excess of cations. This means that the DNA can sometimes not be identified properly. However, there will not be any other new DNA fragments added so no one will be wrongly convicted. Total / Maximum Marks: / 6 marks