Biology 30 Lab Investigation: DNA Extraction from Wheat Germ Name: ______________ Class: _______ Date: _______ Would you like to see LOTS of DNA? This method yields large quantities of DNA that can easily be collected. Instructions 1. Place 1 gram or 1 teaspoon of raw wheat germ in a 50 ml test tube, beaker or jar. 2. Add 20 ml or 1 tablespoon of hot (50-60 °C) tap water and mix constantly for 3 minutes. 3. Add 1 ml of detergent and mix gently every minute for 5 minutes. Try not to create foam! 4. Use an eyedropper, pipette, or piece of paper towel to remove any foam from the top of the solution. 5. Tilt the test tube, beaker or jar at an angle. SLOWLY pour 14 ml of alcohol down the side so that it forms a layer on top of the water/wheat germ/detergent solution. Do not mix the two layers together! DNA precipitates at the water-alcohol interface (the boundary between the water and the alcohol). Therefore, it is crucial to pour the alcohol very slowly so that it forms a layer on top of the water solution. If the alcohol mixes with the water, it will become too dilute and the DNA will not precipitate. 6. Let the test tube, beaker or jar sit for a few minutes. White, stringy, filmy DNA will begin to appear where the water and alcohol meet. You will usually see DNA precipitating from the solution at the water-alcohol interface as soon as you pour in the alcohol. If you let the preparation sit for 15 minutes or so, the DNA will float to the top of the alcohol. You can usually get more DNA to precipitate from the solution by using one of the DNA-collecting tools (such as a glass or paper clip hook) to gently lift the water solution up into the alcohol. This allows more DNA to come in contact with the alcohol and precipitate. You may find it helpful to pour the water/detergent solution into a clean test tube, leaving behind the wheat germ, before adding the alcohol. 7. Use a paper clip hook or a wooden stick to collect the DNA. 8. If you want to keep the DNA, air-dry it on paper towels or filter paper. How Does It Work? Wheat germ This is the DNA source in this protocol. Wheat germ comes from wheat seeds. The "germ" is the embryo, which is the part of the seed that can grow into a new wheat plant. When wheat seeds are milled into white flour, the wheat germ and wheat bran are removed, leaving only starch. Wheat germ contains many nutrients while wheat bran consists of fibre. Whole wheat flour contains all parts of the wheat seed and is therefore more nutritious than white flour while also providing important fibre for digestion. Water temperature The heat softens the phospholipids (fats) in the membranes that surround the cell and the nucleus. It also inactivates (denatures) the deoxyribonuclease enzymes (DNase) which, if present, would cut the DNA into such small fragments that it would not be visible. Denatured enzymes and DNA unravel, lose their shape, and thus become inactive. Enzymes denature at 60° Celsius, and DNA denatures at 80° Celsius. Detergent Detergent contains sodium laurel sulfate, which cleans dishes by removing fats and proteins. It acts the same way in the DNA extraction protocol, pulling apart the fats (lipids) and proteins that make up the membranes surrounding the cell and nucleus. Once these membranes are broken apart, the DNA is released from the cell. Alcohol The DNA released from the cell nucleus is dissolved in the water/detergent/wheat germ solution and cannot be seen. DNA precipitates out of solution in alcohol, where it can be seen. Besides allowing us to see the DNA, the alcohol separates the DNA from the other cell components, which are left behind in the water solution. Answer the following questions on a separate sheet of paper. 1. DNA is the molecule that is passed on from parent to offspring through the mechanism of inheritance. a) What does DNA stand for? b) Describe the shape of DNA. c) What is a nucleotide? d) What is a chromosome? e) What is a gene? f) What is an allele? g) Use a diagram to show the relationship between genes, chromosomes and DNA. 2. DNA is often referred to as the “blueprint for life”. a) In what way can DNA be considered a blueprint? b) What do the A, T, G and C stand for in the genetic code? c) What differentiates one DNA molecule from another? d) If the sequence on one side of a DNA ladder is ATGCCTTGTAAC, what would the opposing side be? e) Critical thinking: Show how you think the following DNA molecule might make an exact copy of itself (in other words, how do you go from one piece of DNA to two identical pieces?). 3. During the late 1990s and early 2000, scientists from around the world worked to figure out the sequence of the human genome (all of the DNA code needed to make a human being). Aptly called the “Human Genome Sequencing Project”, it is considered one of the biggest milestones in scientific history. What type of information could this research provide?