WET LAB DNA Barcoding: From Samples to Sequences - Northwest ...

WET LAB
Lab 3: Analyzing PCR Results with Agarose Gel Electrophoresis
Agarose
Recommended: Agarose, LE Molecular Biology Grade. Available from Hardy
Diagnostics. Item #C8740 (100 gm) or #C8741 (500 gm)
http://www.hardydiagnostics.com
6X DNA loading dye
Recommended: 6X DNA Loading Dye, 5 x 1 ml. Available from Fisher Scientific.
Item # FERR0611, http://www.fishersci.com/
[Note: Some molecular weight standards include free 6X loading dye (see below). In
addition, Lab 3 has been designed for use with classroom micropipettes that adjust in
5 ml increments, requiring 6X loading dye to be diluted to 3X, as described in Teacher
Resource—Aliquoting DNA Barcoding Reagents for Labs 1–4.]
Quantity
Approximately 500 mg
per 2 students
(i.e., 2 students per gel)
2.5 μl per student
Using Bioinformatics: Genetic Research
DNA molecular weight standard
Recommended: GeneRuler 1 kb Plus DNA Ladder, ready-to-use. Available from
Fisher Scientific. Item #FERSM1334, http://www.fishersci.com/
1X Tris Acetate EDTA (TAE) Buffer (“DNA Gel Buffer”)
Recommended: 50X TAE Buffer, 1 L. Available from Fisher Scientific. Item #FERB49
http://www.fishersci.com/
[Note: Dilute 50X TEA buffer to 1X with deionized or distilled water before use.]
Nontoxic DNA gel stain
Recommended: Fast Blast DNA Stain, 100 ml. Available from Bio-Rad.
Item #166-0402EDU, http://www.bio-rad.com/
[Note: Fast Blast DNA Stain should be prepared with deionized or distilled water.]
Class set of Student Handout—Analyzing PCR Results with Agarose Gel Electrophoresis
55°C Water bath or incubator
[Note: Not necessary, but helpful to cool agarose to pouring temperature if desired.]
DNA gel boxes with casting stands and combs
Power supplies
[Note: Most power supplies can power up to two DNA gel boxes.]
Erlenmeyer flasks or glass bottles for melting agarose
15 μl per 2 students
(i.e., 15 μl per gel)
Approximately 50–100 ml
per 2 students
(i.e., 50–100 ml per gel)
Approximately 50-100 ml
per 2 students
(i.e., 50– 100 ml per gel)
1 per student (class set)
1
1 per group
1 per 2 groups
1 per 2 students
(i.e., 1 per gel)
Microwave for melting agarose 1
Hot pads for handling hot agarose
1 per group
1.7 ml microfuge tubes 1 per student
Light Box
[Note: Not necessary, but highly recommended for visualizing DNA gels stained with
Fast Blast DNA Stain or other non-toxic stain.]
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©Northwest Association for Biomedical Research—Updated October 2012