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Protocol of the DNA Extraction in Kitchen

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DNA Extraction's Experiment in Kitchen! This experiment uses kitchen's items only and so easy to do it. You can do the DNA Extraction in your Kitchen!

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Protocol of the DNA Extraction in Kitchen

  1. 1. ©SIProp Project, 2006-2008 1 Protocol of the DNA Extraction in Kitchen Noritsuna Imamura noritsuna@siprop.org
  2. 2. ©SIProp Project, 2006-2008 2 What’s DNA? DNA is a Blueprint for Living Being(Ex.Human). Weight: 0.003ng = 0.000000000003g Chromosome is a Packet for DNA. Human has 46 Chromosomes(2 Sets of 23 Kinds of Chromosome). Nucleus is a Core of Cell. Cell is minimum Unit of Living Being. Human has 37 trillion(37,000,000,000,000) Cells. Source: http://www.ashg.org/education/everyone_1.shtml
  3. 3. ©SIProp Project, 2006-2008 3 Let’s Extract DNA from Banana
  4. 4. ©SIProp Project, 2006-2008 4 Summary Total Time: 20-30mins Protocol 1. Make 10% Saline of 30mL. 2. Cut Banana to 20g. 3. Put Saline, Banana, Dishwashing liquid(6 drops) & Contact lens cleaning solution(6 drops) in Plastic Bag. 4. Mash Banana & Mix them in Plastic Bag. (3mins) 5. Filter “4” Solution in Test Tube. (3mins) 6. Make a micro Tube of DNA Solution 7. Mix 95% Ethanol & Filtered Solution in Test Tube. 1. Pick up Banana’s DNA from Test Tube.
  5. 5. ©SIProp Project, 2006-2008 5 Check Tools Pipette Pipette Chip Dishwash ing Liquid Plastic Bag Scales Beaker Test Tube Micro Tube Tube Stand Funnel & Filter Paper Lens cleaning solution Ethanol Salt
  6. 6. ©SIProp Project, 2006-2008 6 1-1, Make 10% Saline of 30mL Required Tools: Scales Beakers Spoon Muddler Salt CAUTION: Please Skip this part & Use Distilled Water(30mL), if you do the DNA Test Experiment.
  7. 7. ©SIProp Project, 2006-2008 7 1-2, Scale Salt & Water What is “10% Saline of 30mL”? 10% Saline = 90% Water + 10% Salt 30mL = 90% Water + 10% Salt = 27mL Water + 3g Salt Today’s Experiment is Not a Strict Experiment. ≒ 30mL Water + 5g Salt Because it is easy to scale them!
  8. 8. ©SIProp Project, 2006-2008 8 1-3, Mix Salt & Water Use a Plastic Beaker & a Muddler Target Transparency → → →
  9. 9. ©SIProp Project, 2006-2008 9 2-1, Cut Banana to 20g. Required Tools: Scales Cutting Board Knife Banana
  10. 10. ©SIProp Project, 2006-2008 10 2-2, Cut Banana to 20g. Peel & Cut(Slice) Banana. Point: Cut Thin!
  11. 11. ©SIProp Project, 2006-2008 11 3-1, Put All Items in the Plastic Bag Mixed Items: Saline: 30mL Cut Banana: 20g Dishwashing liquid: 6 Drops Contact lens cleaning solution: 6 Drops Required Tools: Plastic Bag
  12. 12. ©SIProp Project, 2006-2008 12 3-2, Put All Items in the Plastic Bag 1. Put Cut Banana 2. Put Saline 3. Drop 6 Drops of Dishwashing liquid 4. Drop 6 Drops of Contact lens cleaning solution
  13. 13. ©SIProp Project, 2006-2008 13 3-3, Close the Plastic Bag
  14. 14. ©SIProp Project, 2006-2008 14 4-1, Mash Banana & Mix them in Bag. Required Tools: Your Hands!
  15. 15. ©SIProp Project, 2006-2008 15 4-2, Mash Banana Mash Banana for 3mins
  16. 16. ©SIProp Project, 2006-2008 16 5-1, Filter “4” Solution in Test Tube Required Tools: Funnel Filter Paper Test Tube Tube Stand
  17. 17. ©SIProp Project, 2006-2008 17 5-2, Fold Filter Paper 1. Fold Crescent-shape 2. Fold Quadrant of a Circle 3. Make Cone-shape (Open the Filter Paper)
  18. 18. ©SIProp Project, 2006-2008 18 Easy Way of Filtering Mashed Something Use Gauze Sheets as 1st Filter & a Coffee Filter as 2nd Filter. Gauze(1st Filter) prevents clogging of a Filter Pater/Coffee Filter with Mashed Banana. A Coffee Filter is easier to buy a Paper Filter.
  19. 19. ©SIProp Project, 2006-2008 19 5-3, Set Up Funnel & Test Tube 1. Set The Cone in the Funnel 2. Set the Test Tube into the Tube Stand 3. Insert the Funnel to the Test Tube
  20. 20. ©SIProp Project, 2006-2008 20 5-4, Drip the Mashed Banana Solution 1. Move the Solution to the One Side of the Bag 2. Cut the Bag of Opposite Side 3. Pour the Solution into the Funnel 4. Wait for 3mins
  21. 21. ©SIProp Project, 2006-2008 21 5-5, Check the Amount of Solution Pick up the Funnel & the Test Tube together Check the Amount of Filtered Solution DNA melts into this solution. Minimum Solution
  22. 22. ©SIProp Project, 2006-2008 22 Try to Make one more Tube!
  23. 23. ©SIProp Project, 2006-2008 23 6-1, Make a micro Tube of DNA Solution Required Tools: Micro Pipette (200uL or 1000uL) Pipette Chips Micro Tube (0.2mL=200uL) aka PCR Tube
  24. 24. ©SIProp Project, 2006-2008 24 6-2, Set Pipette’s Amount We need “100uL” Liquid. Set Scale Dial at “100(200uL)” or “010(1000uL)” Amount Dial
  25. 25. ©SIProp Project, 2006-2008 25 6-3, Set Up Pipette Chip Insert Pipette Chip to Tip of the Pipette
  26. 26. ©SIProp Project, 2006-2008 26 6-4, How to Use the Pipette? Push Button has 2 kinds of type. 1st Stop・・・Suck Operation 2nd Stop・・・Flash Operation Procedure 1. Push & Keep the Push Button to 1st Stop 2. Insert the Pipette Chip into the Solution 3. Release the Push Button Slowly 1. Suck the Solution with the Pipette. 4. Extract the Pipette from the Test Tube 5. Insert the Pipette Chip into a micro Tube 6. Push & Keep the Push Button to 1st Stop 1. & Keep this position 1sec 7. Push & Keep the Push Button to 2nd Stop(Flash) 8. Extract the Pipette from the micro Tube Push Button
  27. 27. ©SIProp Project, 2006-2008 27 6-5, Move the Solution to a micro Tube Move the Solution from the Test Tube to a micro Tube Suck the Filtered Solution from the Test Tube Procedure 1. Push & Keep the Push Button to 1st Stop 2. Insert the Pipette Chip into the Solution of the Test Tube 3. Release the Push Button Slowly 1. Suck the Filtered Solution with the Pipette. 4. Extract the Pipette from the Test Tube
  28. 28. ©SIProp Project, 2006-2008 28 6-6, Move the Solution to a micro Tube Move the Solution from the Test Tube to a micro Tube Move the Filtered Solution to the micro Tube Procedure 5. Insert the Pipette Chip into a micro Tube 6. Push & Keep the Push Button to 1st Stop 1. & Keep this position 1sec 7. Push & Keep the Push Button to 2nd Stop(Flash) 8. Extract the Pipette from the micro Tube
  29. 29. ©SIProp Project, 2006-2008 29 6-7, Throw the Pipette Chip Push the Eject Pipette Chip Button on Garbage. Please Change the Pipette Chip by each Using Pipette. Eject Pipette Chip Button
  30. 30. ©SIProp Project, 2006-2008 30 6-8, Why made it? This Tube is used in DNA Test Phase. For PCR(Polymerase Chain Reaction) Sequence Amplify a single copy or a few copies of a piece of DNA. https://en.wikipedia.org/wiki/Polymerase_chain_reaction
  31. 31. ©SIProp Project, 2006-2008 31 Please Stop this Experiment, if you do the DNA Test Experiment. And Go to “Protocol of the DNA Test in Kitchen”
  32. 32. ©SIProp Project, 2006-2008 32 7-1, Mix Ethanol & the Filtered Solution Required Tools: 90% over Ethanol (1-2 Times Amount of the Filtered Solution) The Tubes of the Filtered Solution The Test Tube The micro Tube
  33. 33. ©SIProp Project, 2006-2008 33 7-2, Put Ethanol into the Test Tube Using the Funnel without a Filter 1-2 Times Amount of the Filtered Solution
  34. 34. ©SIProp Project, 2006-2008 34 7-3, Pick Up DNA 1. Insert a Fork into the Test Tube 2. Stir this Solution Super Slowly by the Fork 1. A White Haze gradually appears. 3. Pick Up this White Haze 1. This is Banana’s DNA!!!
  35. 35. ©SIProp Project, 2006-2008 35 7-4, The micro Tube Side Mix Ethanol(100uL) Put Ethanol into the micro Tube by Pipette
  36. 36. ©SIProp Project, 2006-2008 36 7-5, Flick the micro Tube with a Finger When Flick a finger, A White Haze appears. This is called “Tapping Method” in Bio Science
  37. 37. ©SIProp Project, 2006-2008 37 The Principle of This Experiment
  38. 38. ©SIProp Project, 2006-2008 38 The Principle of Surfactant Dishwashing Liquid This alias is Surfactant. This is a catalyst which can mix oil with water. Surfactant https://en.wikipedia.org/wiki/Surfactant Cell & Cell nuclear have membrane. A structure of Cell Membrane & Nuclear Membrane is Lipid(≓oil) Bilayer. Lipid Bilayer https://en.wikipedia.org/wiki/Lipid_bilayer Dishwashing Liquid can break down Cell Membrane & Nuclear Membrane. Source: https://en.wikipedia.org/wiki/Cell_(biology)#/media/File:Wilson1900Fig2.jpg
  39. 39. ©SIProp Project, 2006-2008 39 The Principle of Protease Contact lens cleaning solution One of the ingredients of this is protein-degrading enzymes(=Protease). Protease https://en.wikipedia.org/wiki/Protease Chromosome is made by DNA & some kinds of Proteins. Contact lens cleaning solution can separate Chromosome into DNA & some kinds of Proteins.
  40. 40. ©SIProp Project, 2006-2008 40 The Principle of Filter Filter DNA is called Nucleic Acids. Nucleic Acids melts into water. Your Hand Surfactant Protease Water
  41. 41. ©SIProp Project, 2006-2008 41 The Principle of Ionization Saline(Salt) Salt alias is Sodium Chloride. Chemical Formula of Sodium Chloride: NaCl Saline alias is Electrolyte Solution of NaCl(= NaCl is ionized to Na+ & Cl- in water) Ionic Formula of Sodium Chloride: NaCl→ Na+ + Cl- DNA Ionization One part of the DNA backbone is a phosphate group. Red Circle Part is ionized to P-O-(Ionized DNA) & H+ in water. Ionic Formula : DNA→ H+ + P-O-(Ionized DNA) DNA Ionization Status is Negative(-). Repulsive force acts on each Ionized DNA. Ionized DNA can NOT fit a lump. Source: http://www.chemguide.co.uk/organicprops/aminoacids/dna1.html#top
  42. 42. ©SIProp Project, 2006-2008 42 The Principle of Ionic Bond Ionic Bond Between Positive(+) Ion & Negative(-) Ion bond. Positive(+): Na+ from Salt Negative(-): P-O-(Ionized DNA) from DNA Ionic Formula : Na+ + P-O-(Ionized DNA) → P-ONa = DNA Natrium DNA Natrium doesn’t have Electric Charge. DNA Natrium CAN fit a lump.
  43. 43. ©SIProp Project, 2006-2008 43 The Principle of Precipitation 95% Ethanol When Solution of the Filtered Solution & Ethanol are mixed, DNA Natrium moves the Filtered Solution(Water) side to Ethanol side. Solubility: the Filtered Solution(Water) > Ethanol DNA Natrium Lump is precipitated in Ethanol.
  44. 44. ©SIProp Project, 2006-2008 44 Next Protocol of the DNA Test in Kitchen

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