DNA Fingerprinting

Supplies from Iowa State

Student Preparation

DNA and Enzyme Preparation

Obtaining Supplies(Iowa Teachers Only)

Teachers should request the following supplies (next slide) at least a week in advance.

Supplies should be ordered by contacting Lori Miller by phone in Iowa, 1/800-643-9504 or 515/294-9818,

or by Fax: 515/294-4629.

The electrophoresis equipment can be obtained by contacting the Area Education Agency.

DNA Fingerprinting

Supply List 6 boxes of pipette tips (already autoclaved)

34 µl of 0.025 µg/µl pBR322 per class section (minimum of 68 ml per school)

17 µl of 0.075 µg/µl Ad-2 DNA per class section (minimum of 34 ml per school)

17 µl of 0.15 µg/µl l DNA per class section (minimum of 34 ml per school)

17 µl of 0.025 µg/µl pUC19 DNA per class section (minimum of 34 ml per school)

3 µl Bgl 1 per class section (minimum of 6 ml per school)

15 µl reaction buffer per class section (minimum of 30 ml per school)

DNA Fingerprinting

Supply List(cont.)

144 µl Carolina Blu Gel Stain per gel (minimum of 2 gels)732 µl Carolina Blu Buffer Stain per gel (minimum of 2 gels)

1 bottle Carolina Blu DNA Stain 0.7 g agarose per gel (minimum of 2 gels)

70 ml 10X TBE per gel (minimum of 2 gels)40 ml blue migration dye for each student group8 microcentrifuge tubes for each student group

Fingerprinting instructions for each student

*Please send the unused pipette tips/boxes, 10X TBE bottles (rinsed out) and Carolina Blu DNA Stain

back to the Office of Biotechnology when finished.

DNA Fingerprinting

Preparation of the Student Materials

• The supplies can best be provided to the class in groups of five students.

• The DNA samples should be refrigerated until class time.• It is too expensive for the Office of Biotechnology to

provide DNA for every student in a class. The office will provide enough DNA, restriction endonuclease, and reaction buffer for a minimum of two groups of five students or a maximum of one group of five students in every class section, whichever is greater.

DNA Fingerprinting

DNA Fingerprinting Preparation of the Student Materials

(cont.)

• For the other student groups, use distilled water to replace the DNA, restriction endonuclease, and reaction buffer.

• Every group of students should be provided with the blue migration dye.

DNA Fingerprinting

DNA Fingerprinting

DNA and Enzyme

Lab Preparation

DNA and Enzyme PrepStep 1

• For each group of students label 7-1.5 ml tubes "C,1,2,3,4, N and D" with a permanent felt pen.

• Tube "N" will contain the enzyme mixture.

• Tube "D" will contain the migration dye.

DNA Fingerprinting

DNA and Enzyme PrepStep 2

• Pipette 17 µl of a 0.025 µg/µl concentration of pBR322 DNA into the tube labeled "C."

DNA Fingerprinting

DNA and Enzyme PrepStep 3

• In the tube labeled "1" pipette 17 µl of 0.15 µg/µl concentration of lambda DNA.

DNA Fingerprinting

DNA and Enzyme PrepStep 4

• In the tube labeled "2" pipette 17 µl of 0.075 µg/µl concentration of Ad-2 DNA.

DNA Fingerprinting

DNA and Enzyme PrepStep 5

• In the tube labeled "3" pipette 17 µl of 0.025 mg/ml concentration of pBR322 DNA.

DNA Fingerprinting

DNA and Enzyme PrepStep 6

• In the tube labeled "4" pipette 17 µl of 0.025 µg/µl concentration of pUC19 DNA.

DNA Fingerprinting

DNA and Enzyme PrepStep 7

• In the microcentrifuge tube (1.5 ml) labeled "N," pipette 15 µl of reaction buffer and 3 µl Bgl 1.

DNA Fingerprinting

DNA and Enzyme PrepStep 8

• In a microcentrifuge tube (1.5 ml) labeled "D," pipette 40 µl of blue migration dye.

DNA Fingerprinting