19.3: Pull Sheets/Materials

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Page ID: 118044

Darcy Ernst, May Chen, Katie Foltz, and Bridget Greuel
Evergreeen Valley College via Open Educational Resource Initiative at Evergreen Valley College

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The following DNA sample and restriction endonucleases will need to be ordered and arrive/stored prior to the start of this lab:

lambda DNA (bacteriophage lambda, 48,502bp) 200uL (100uL/section)
- Thermofisher Catalog # SD 0011 (or equivalent)
  - https://www.thermofisher.com/order/catalog/product/SD0011
  - $154 for one (one should be enough for both lab sections)
  - Note: volumes are based on 0.3 ug/uL concentration

Pst I enzyme – keep on ice 12uL (6uL/section)
- Need 6uL per section
  - https://www.thermofisher.com/order/catalog/product/ER0611
  - $28.98 for one (one should be enough for both lab sections)
  - Should come with buffer (need 30 uL of buffer /section)

Eco RI enzyme - keep on ice 12uL (6uL/section)
- Need 6uL per section
  - https://www.thermofisher.com/order/catalog/product/ER0271
  - $47.38 for one (one should be enough for both lab sections)
  - Should come with buffer (need 30 uL of buffer /section)

Hind III enzyme – keep on ice 12uL (6uL/section)
- Need 6uL per section
  - https://www.thermofisher.com/order/catalog/product/ER0501
  - $39.09 for one (one should be enough for both lab sections)
  - Should come with buffer (need 30 uL of buffer /section)

PREPARED MATERIALS:

Item	How many?
Gloves (s, m, l)	1 box of each
Electrophoresis chambers and power boxes models with dials/scales to adjust voltage	6
Electrophoresis trays and combs	6
Pans or trays (clean) To hold electrophoresis chambers	6
Ice bucket with ice	2
Extra Microcentrifuge (one is already in the lab room, need 2 total)	1
P-20 micropipettes	12
Micropipette tips for P-20	6 boxes
P-2 micropipettes	12
Micropipette tips for P-2	6 boxes
Plastic cling wrap	1 roll
Sharpie™ pens	6
Large beaker or jar labeled – “Used Pipette Tips and DNA Vials”	3 (1 per bench)
Waste container for methylene blue stain (keep in safety cabinet/hood)	1
Microtubes for practice	14 (7 per lab section)
Microtube racks	6
Water bath set to 55-60°C (for agarose)	1
Water bath set to 37°C (for enzyme digest)	1
7x7 plastic (Tupperware) boxes with lids for staining	12 (6/section)
Practice gels covered with water in Petri dishes	14 (7/section)
Agarose gel solution in flask (keep in 55°C water bath)	14 (7/section)
1X TAE solution Divided into 2-3 flasks that students can pour (or enough for 7 gels and afternoon lab can reuse the morning lab solution)	enough for 14 groups in total
Practice solution – Colored Glycerin Water 1 mL in each microtube	14 total microtubes
0.025% Methylene Blue Staining Solution	enough for 14 gels
Microtubes with gel loading dye At least 15uL per microtube Do not dilute loading dye (keep it as it comes, 6x)	14 microtubes
4 color-coded microtubes (keep empty, students will fill) Labeled: L, P, E and H (different color for each tube)	12 of each color (6 of each per lab section)

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