K-12 Programs

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PCR Field Trip
Set-up

Materials/Equipment Needed

Generally the set up is for 4 students per lab bench.

I. Equipment needed on each lab bench where students will work:

• 4 PCR protocols (these should not be in sleeves, nor written on)
• 4 goggles
• 4 lab coats
• 1 waste beaker
• 1 box of Kimwipes
• 1 ice bucket
• 1 table top microcentrifuge
• 2 p20 Pipetman
• 2 p200 Pipetman
• 1 p1000 Pipetman
• 2 yellow tip boxes
• 1 blue tip box
• 2 blotting pads (for pipetting practice)
• 2 tubes of colored dyes (for pipetting practice)
• 1 bottle of nuclease free water
• 1 dropper bottle of mineral oil
• 1 50ml conical tube containing 5 0.5ml clear Ep. tubes
• 1 test tube rack containing:

  4 Sharpie pens (with at least 1 or 2 fine-point)
  5 clear thin-walled 0.5ml PCR tubes (found in PCR drawer);
  1 0.5ml blue Ep. Tube (for Master Mix) 
  2 tubes of 6X blue/orange loading dye

II. Equipment needed on lab benches where students will run gels:

• 1 gel box for every bench/group (4 students)
• 1 power supply for every 2 gel boxes
• O.5X TBE in a jug, about 1 L per 2 gel boxes
• 1 125 or 150ml flask containing premelted 2% agarose (1 g agarose in 50mls 0.5X TBE)
• 1 6 well gel comb for each group
• 1 gel tray for each group
• tape to tape gel trays, if necessary
• 1 box Kimwipes

III. Reagents/Equipment controlled by Instructor

• Ethidium bromid
• Thermocycler in lab 232 (Reserved ahead of time for the lab)
• 1 tube of Template DNA per student for PCR reactions.

The aliquots are stored in a purple test tube rack, and the stock tubes are stored in a Styrofoam box, in the refrigerator in 233. There should be a variety of DNA sample numbers in a variety of 0.5ml colored Ep. Tubes.

Each aliquot contains 3ul of template DNA (100ng/ul).

IV. Reagents needed in each ice bucket (1 bucket per bench)

 

Stocks	Quantity	Reagent	0.5ml Ep tube color	Aliquot	Storage
100ml aliquots in PCR FT box @ -20°C	1	F Forward Primer (100 pmol/ml )	green	8 ul	-20°C (or 4°C)
100ml aliquots in PCR FT box @ -20°C	1	R Reverse Primer (100 pmol/ml )	blue	8 ul	-20°C (or 4°C)
Promega product tube in PCR FT box @ -20°C	1	10x Taq PCR buffer	orange	40 ul	4°C (or-20 °C)
Promega product tube in PCR FT box @ -20°C	1	Taq Pol (5 u/ml )	pink	2 ul	-20°C
Promega product tube in PCR FT box @ -20°C	1	dNTP (10mM total, 2.5 mM each nucleotide)	yellow	10 ul	-20°C
PCR FT box @ 4°C	1	100 bp marker* (130 ng/ml )	clear	8 ul	4°C

• 100 bp marker: (catalog # G2101). Prepare 2:3:1 ratio as follows:
  250 ul marker in new tube: 375 ul ddH2O: 125 ul 6X dye
  (2 ul marker: 3 ml ddH2O: 1 ul 6X dye)

• Background
• Genetic Screening Scenario
• Information for Teachers
• Protocol
• Biotechnology Field Trips Home Page

Contact: Barbara Bielec, 608-273-9737