DNA EXTRACTION

DNA EXTRACTION USING STRAWBERRIES Activity Overview Extract DNA from strawberries using simple household chemicals

Questions: what does DNA stand for? what is DNA?

DNA - Deoxyribonucleic acid! All living organisms contain DNA in their cells – FOUND in nucleus. DNA - Deoxyribonucleic acid! The nucleus is a membrane bound structure that contains the cell's hereditary information and controls the cell's growth and reproduction. It is commonly the most prominent organelle in the cell. You mean I can see it? HOW? www..kamibudaksains.blogspot.com DNA - The complex chemical compound found in chromosomes that contains the genetic code.

Figure: http://employees. csbsju Figure: http://employees.csbsju.edu/hjakubowski/classes/ch331/dna/oldnastructure.html Packaging of DNA in the Nucleus

Do you think you have very much in common with a strawberry Do you think you have very much in common with a strawberry? Believe it or not, a strawberry’s genetic material is very similar to your own! Today you will see and touch dna.

New words Add these to your notebook MEANING: Extraction Precipitate Spooling Denaturising Lysing Soluble Define these words at the end of the lesson

Set up your lab notebook Title: EXTRACTING DNA FROM STRAWBERRIES Introduction: Why do we need to learn about DNA? Why is DNA important to living things? What does DNA look like? Purpose To isolate and visualize DNA from a strawberry Hypothesis DNA from strawberries looks like ...

MATERIALS (per two students) 1 Ziploc bag 1 strawberry 10 ml DNA extraction buffer (soapy, salty water) Meat tenderizer 1 piece of cheesecloth Funnel Glass rod or wooden stick Ice-cold isopropyl alcohol Test tube Test tube rack Parafilm

METHOD Form groups of two students. Student #1 pick up a Ziploc bag and a strawberry, a piece of cheese cloth, funnel, and glass test tube, parafilm and wooden stick. Put the strawberry in the Ziploc bag and squish it for two minutes Question: Why are you squishing the strawberry here? What is happening to the cells in the strawberry?

METHOD: Student #2 pour the 10 ml of cold extraction buffer in the test tube into the bag, then smash the strawberry around for one minute or more until no big chunky parts remain. Wait three minutes. Describe your observations of the strawberry? What is happening to the cells after you added the extraction buffer (salt and detergent)?

What is the extraction buffer? SALT DISHWASHING DETERGENT

METHOD: Student #1 pour the strawberry mush through a funnel lined with cheesecloth. Allow the fluid to collect in a glass test tub until the test tube is about 1/3th of the way full. Add a small pinch of meat tenderizer. Parafilm the tube and Mix GENTLY. Why is cheesecloth used here to help you extract DNA from the strawberry?

WHAT MUST I DO NOW? MEAT TENDERIZER ALCOHOL SEPARATION ENZYME POWDER www.forums.overclockers.com.au

METHOD: Student #2 retrieve isopropyl alcohol from your teacher. While holding the test tube at an angle, gently and SLOWLY trickle the cold isopropyl alcohol down the side of the tube until it is half full. DO NOT shake the tube!

Allow test tube to sit for several minutes. METHOD: Observe the mixture for a few minutes. You will see a white, threadlike substance rise from the mixture to rest above in the alcohol layer. This is the DNA that you have extracted from the cells of the strawberry. Allow test tube to sit for several minutes. The clearer the DNA is the fewer impurities you have. Draw what you see in the test tube. Identify and label all the layers. Be sure to note color, relative volume, and thickness of materials in each layer. Which layer has the white precipitate?

WOW – SPOOLING THE DNA!!! Student #1 dip the wooden stick into the tube where the alcohol and strawberry layers meet. Gently twist the stick and try to catch some on it. Do not stir, and don’t go too quickly. Sometimes, the clear fibrous DNA may float to the top of the liquid. These are not single DNA molecules, but huge ropes of thousands of molecules twisted together. Gently lift the water solution up into the alcohol layer (this allows more DNA to get in contact with the alcohol and precipitate).

Strawberry DNA Describe what you see as you try to lift the white precipitate (DNA) out of the tube. What does it look like? What does it feel like? It is easy to grab? What color is it?

CLEAN UP! Toss the cheesecloth and bags into the garbage can. Rinse your test tubes and place them on the rack to dry Clean up your area and put supplies back where you found them Wash your hands Return to your desk and answer the questions.

WHAT IS THAT STRINGY STUFF. DNA is a long, stringy molecule WHAT IS THAT STRINGY STUFF? DNA is a long, stringy molecule. The salt that you added in step one helps it stick together. So what you see are clumps of tangled DNA molecules!

DNA normally stays dissolved in water, but when salty DNA comes in contact with alcohol it becomes undissolved. This is called precipitation. The physical force of the DNA clumping together as it precipitates pulls more strands along with it as it rises into the alcohol.

THE WHY’S!!!! Squishing separated the strawberry cells. In order to extract DNA from a cell, the associated membranes and proteins must first be removed and then physically separated from the DNA. Salt and Detergents are used to break down cell walls and nuclear membranes to release the DNA. They work by chemically poking holes in the cell membranes or walls. Once holes are poked in the membranes, the membranes can be further disrupted mechanically, (you squished it) After that, it is easier to get the contents of the cell out, including the DNA. Sodium is an element. It is a positive ion and often associates with negative ions as part of useful compounds. Salt helps strip away the proteins associated with DNA. + and – charge

THE WHY’S!!!! Why detergent? Each cell is surrounded by a sack (the cell membrane) DNA is found inside the second sack (nucleus) within each cell. To see the DNA we have to break it open. A cell membrane has 2 layers of lipid(fat) molecules with protein going through them. When the lysis buffer (detergent) comes close to the cell, it captures the lipids and the proteins - breaks open the cell destroying the fatty membrane - DNA now released into the solution.

A CELL'S MEMBRANES HAVE TWO LAYERS OF LIPID (FAT) MOLECULES WITH PROTEINS GOING THROUGH THEM.

Why detergent. How does detergent work Why detergent? How does detergent work? Think about why you use soap to wash dishes or your hands. To remove grease and dirt, right? Soap molecules and grease molecules are made of two parts: 1.(Blue) Heads, which like water. 2.(Green) Tails, which hate water.

After adding the detergent, what do you have in your soup?

THE WHY’S!!!! enzyme (meat tenderizer) The DNA in the nucleus of the cell is molded, folded, and protected by proteins. The tenderizer cut the proteins away from the DNA  In this experiment, meat tenderizer acts as an enzyme to cut proteins just like a pair of scissors. The meat tenderizer cuts the proteins away from the DNA

THE WHY’S!!!!alcohol Alcohol is less dense than water, so it floats on top. DNA is not soluble in alcohol - other cell parts are. DNA precipitates out of the solution and collect at the interface of the alcohol and soap layer. The colder the alcohol the less soluble the DNA will be in it. Colder temperatures slow down enzymes that can break down DNA, giving better extraction results.

Why Is DNA Extraction Important? DNA extraction is an important molecular biology procedure. By definition, extraction is taking DNA out of any type of cell for the purpose of analysis.

PART B: OBSERVING EXTRACTED DNA UNDER THE MICROSCOPE

PART B: A CLOSER LOOK MATERIALS / EQUIPMENT ➤ Strawberry DNA ➤ Dropping pipet ➤ methylene blue ➤ microscope ➤inoculation needle ➤ microscope slide and cover slip ➤ paper towelling 31

PART B: A CLOSER LOOK Using a dropping pipette, carefully remove some of the threadlike substance from the top of your preparation. Place one or two drops onto the middle of a microscope slide. Add two drops of methylene blue. Wait 3 or 4 minutes to allow the methylene blue to be absorbed by the DNA. Carefully place a cover slip on the slide. Gently press a folded piece of paper towelling over the top of the prepared slide to soak up any excess liquid. Observe the DNA under low power, then high power. 

DISCUSSION Answer in group related work. 1 Write a detailed description of the material floating at the top of the test tube after the alcohol was added.   2 Describe the DNA as it appears under the microscope under high power. 3 Prepare a diagram of your DNA specimen. 4 What was the reason for using methylene blue?

Construct a flow chart / drawing/ mind map of the process you used extracting the dna from Strawberries Next to each step explain how the method you used was important. What substances make up the membranes of cells and cell organelles? Where can DNA be found in the cell? Discuss the action of the soap (detergent) on the cell. What is the purpose of the soap in this activity? What was the purpose of the Sodium Chloride? Include a discussion of polarity and charged particles. What does the meat tenderizer do? Why was the cold ethanol added to the soap and salt mixture? Describe the appearance of your final product?

TIPS: Steps for flow chart In order to release the DNA from the nuclei of the strawberry cells you first separated the cells from one another. Then the cell and nuclear membranes needed to be ruptured to release the cell contents and the contents of the nucleus. Once removed from the nuclear membrane the DNA had to be untangled into the visible, threadlike structures you ended up with.

HOMEWORK: EXTENSION GROUP QUESTIONS Where can DNA be found in the cell? Discuss the action of the soap (detergent) on the cell. What is the purpose of the soap in this activity? What was the purpose of the Sodium Chloride? Include a discussion of polarity and charged particles. Why was the cold ethanol added to the soap and salt mixture? Describe the appearance of your final product? Draw a diagram of DNA containing 5 sets of nucleotide bases labelling the hydrogen bonds between the bases. References and Resources: Adapted from Berry Full of DNA by Diane Sweeney for Biology: Exploring Life to be published by Prentice Hall. Websites: http://www.carlinvilleschools.net/linke/Biology/DNA.htm http://carnegieinstitution.org/first_light_case/horn/DNA/dn aindex.html

YOUR EXTRACTED dna under the microscope looked like .........! Collaboration-Brainstorming on what to look at under the - DNA extraction- DNA is too small for even a microscope to see and after the extraction it just appears like a blob. True or false? Could you any DNA strand? www.employees.csbsju.edu

References: http://www.squidoo.com/how-to-get-dna-from-a-kiwi-fruit Why Is DNA Testing Good? | eHow.com http://www.ehow.com/about_6684410_dna- testing-good_.html#ixzz1MkbcIJs5 Why Is DNA Extraction Important? | eHow.com http://www.ehow.com/list_5839095_dna-extraction-important_.html#ixzz1MkZDrqyY Why Is Sodium Used in DNA Extraction? | eHow.com http://www.ehow.com/about_6504902_sodium-used-dna- extraction_.html#ixzz1MkaGWg57 Why Is Cold Alcohol Used in DNA Tests? | eHow.com http://www.ehow.com/about_6399349_cold-alcohol-used-dna- tests_.html#ixzz1MkbEmEdu http://learn.genetics.utah.edu/content/labs/extraction/howto/  www.kamibudaksains.blogspot.com http://www.chemwatch gold.com References and Resources: Adapted from Berry Full of DNA by Diane Sweeney for Biology: Exploring Life to be published by Prentice Hall. Websites: http://www.carlinvilleschools.net/linke/Biology/DNA.htm http://carnegieinstitution.org/first_light_case/horn/DNA/dnaindex.html

References: Uses of DNA Extraction | eHow.com http://www.ehow.com/about_5344428_uses-dna- extraction.html#ixzz1MkWQB2TZ Science 10: A Contextual Approach Heineman Queensland Science Projects : Regan Spence Maggie Spenceley http://en.wikipedia.org/wiki/Molecular_biology http://www.clinchem.org/cgi/content/extract/44/10 /2201 http://employees.csbsju.edu/hjakubowski/classes/c h331/dna/oldnastructure.html http://www.genomebc.ca/education/teachers/class room-activities/dna-extraction/