Crown Jewels/Get a Clue Educator Outline

* Intro. To Lab – robbery of the royal family in London – OR – ipod – OR – Homecoming Tiarra

-blood samples w/ DNA evidence collected at scene

-2 suspects were elim. (alibi & wrong blood type)

-DNA is unique to each person

-Fragments are diff., Identical twins have identical DNA

-Amplified DNA – PCR (Polymerase Chain Reaction) – repeats DNA sequence over & over

-You are the Forensics Team that has been hired to determine the culprit of this attempted robbery – gown, goggles, gloves

- Materials: DNA sample (kept on ice – heat can break it down) + restriction enzyme (cuts DNA)

+ loading dye (weighs down sample & stains it) + buffer (keeps pH stable)

- STEP #1: Tap and heat DNA (put into heating blocks – heats straightens out the twist allowing it to

move thru- the agarose; releases sticky ends)60-65 C for 3-4min.

Illustration - DNA fingerprinting to introduce gel - load gels – to +, apply electricity

(DNA – & will move to + end) – Finished product is the Fingerprint of that DNA

- small fragments move farther; larger fragments will be closer to wells

Your Equipment:

* Your gel with wells – wells should be at - (black) end of chamber – blue liquid –

**DON’T TOUCH** (DNA movement would be messed up)

* Buffer completely covers gel (conducts electricity)

* Electrophoresis Chamber (- DNA will move to + end)

* Power Supply –______V! (**BE CAREFUL & FOLLOW DIRECTIONS**)

* Digital micropipettes - **Demostrate how to hold & read dial – IF NOT ALREADY DONE**

Illustration- uL symbol (1000uL = 1mL) ***Show Microtube (approx. 1mL)***

Pipette Use: (IF NOT ALREADY DONE)

-Never put pipette into liquid w/o tip

-**Instruct on how to use**

-Pick up & release LD a few times until comfortable

-Pipettes back on rack

Loading Wells: * Camera * - picture (tip just below liquid level but not in well)

-“Pool Technique” (2 hands) w/ support

Practice Loading - IF NOT ALREADY DONE IN PREVIOUS CLASS

**Don’t poke into gel***

Ready to Load real DNA? – Name on paper

  • Illustration * - map of gel - Fill in your paper (*if make a mistake, fix paper, don’t try to take out of gel*) – use a steady hand & LOAD GEL SLOWLY!!!

STEP #2: * get DNA microtubes (C,S1 & S2) out of heating block

  • get new tip and Mix up DNA (draw up & release 2x) – 3rd draw up is the loader

STEP #3:* Load gels (mix before loading each microtube) – Partners take turns loading

  • Guide each other to the correct using your marked paper

STEP #4:* examine lid closely w/holes that match up to posts on chamber

  • **match Red to Red & Black to Black**
  • **put lid on chamber & press firmly until you hear a snap**

STEP #5:* Plug into power supply (wait until both groups finish before turning on – Run button)

  • will run for about ______minutes
  • should see bubbles rising & begin to see DNA moving (purple dye moving)

How to Make Gel:***DEMO IF THIS IS TO BE INCLUDED***

  • Agarose powder (from seaweed) + buffer (has water – w/ acids & bases – stabilizes pH)
  • Vortex (to mix) then heat & cool
  • Carolina Blu (1 drop – for staining)
  • Vortex
  • Casting Tray (acrylic tray – will solidify in tray for shape)
  • Comb makes wells (for DNA to be loaded into) - **Let a students pull out comb**
  • **let students touch gel**

Work on Questions:

Pipette volume instruction: ***IF NOT ALREADY DONE***

-* Camera* - dial numbers & how to read

-red 0 is the decimal place

-show how to change volume

-should be set back on 20 uL (0 – 20uL) **depending on what module is next class**

Volume Chart on back of Data Sheet:

Career Info: Biotech. Encouragment, CSI, NC the 3rd leading state in the nation in BioTech

STEP #6:* Push Stop Button (power supply off)

  • Unplug chambers
  • Remove probs from power supply
  • Take tops off chambers (thumbs push down on posts will help release top)
  • Wrap cords back (Be careful not to get probs in chamber water)

STEP #7:* Carefully lift tray out of chamber, drain excess liquid off,

  • Carefully tilt tray over weigh boat so gel gently slides off into boat

WHO IS THE CULPRIT BASED ON YOUR BANDS?

WHOSE BANDS MATCH THE CRIME SCENE DNA?

HAVE STUDENTS MARK BANDS ON LAB PAPERS!!