Bio-Rad PV92 PCR Informatics Kit User Manual

1. Label one 1.5 ml micro test tube with your
initials. Label one screwcap tube containing
200 µl of InstaGene matrix with your initials.

2. Obtain a cup containing saline solution from
your instructor. Pour the saline into your mouth
and rinse vigorously for 30 seconds. Expel the
saline back into the cup.

3. Transfer 1 ml of your saline rinse into the micro
test tube (NOT the screwcap tube) with your
initials. If a P-1000 micropipet is not available,
carefully pour ~1 ml of your saline rinse into
your micro test tube (use the graduations on the
side of the micro test tube to estimate 1 ml).

4. Spin your tube in a balanced centrifuge at full
speed for 2 minutes. When the centrifuge has
completely stopped, remove your tube. You
should see a match-head sized pellet of whitish
cells at the bottom of the tube. If you don’t see
a pellet of this size, decant the saline, refill your
tube with more of your oral rinse, and repeat the
spin.

5. After pelleting your cells, pour off the saline.
Being careful not to lose your pellet, blot your
tube briefly on a paper towel or tissue. It’s OK
for a small amount of saline (< 50 µl, about the
same size as your pellet) to remain in the
bottom of the tube.

6. Resuspend the pellet by vortexing or flicking the
tube so that no clumps of cells remain.

7. Using a 2–20 µl adjustable-volume micropipet
set to 20 µl, transfer all of your resuspended
cells to the screwcap tube containing
InstaGene.

8. Screw the cap tightly on the tube. Shake or
vortex to mix the tube contents.

33

Quick Guide

Lesson 1 Cheek Cell DNA Template Preparation

Centrifuge

9. When all members of your team have
collected their samples, place the tubes in the
foam micro test-tube holder, and incubate at
56°C for 10 minutes in a water bath. At the
halfway point (5 minutes), shake or vortex the
tubes gently, then place back in the 56°C water
bath for the remaining 5 minutes.

10. Remove the tubes, shake or vortex, and place
the tubes in a boiling water bath (100°C).
Incubate at 100°C for 5 minutes.

11. Remove the tubes from the boiling water bath
and shake or vortex the contents to resuspend.
Pellet the matrix by spinning at 6,000 x g for
5 minutes (or 2,000 x g for 10 minutes) in a
centrifuge.

12. Store your screwcap tube in the refrigerator until
the next laboratory period (or proceed to step 2
of Lesson 2).

34

Centrifuge

56°C, 10 min

100°C, 5 min

Water bath

Water bath

Quick Guide

1. Label 1 screwcap tube containing 200 µl of
InstaGene matrix plus protease with your
initials.

2. Collect 2 hairs from yourself. Select hairs that
have either a noticeable sheath (a coating of
epithelial cells near the base of the hair), or a
good sized root (the bulb-shaped base of the
hair). Trim the hair, leaving the last ~2 cm of the
base of the hair. Place the trimmed hairs into
the screwcap tube with your initials.

3. Place your tube in the foam micro test-tube
holder and incubate at 56°C for 10 minutes in
a water bath. At the halfway point (5 minutes),
shake or vortex gently, then place it back in the
56°C water bath for the remaining 5 minutes.

4. Remove the tubes, gently shake or vortex, and
place them in a boiling water bath (100°C).
Incubate at 100°C for 5 minutes.

5. Remove the tubes from the boiling water bath
and shake or vortex the contents to resuspend.
Pellet the matrix by spinning at 6,000 x g for
5 minutes (or 2,000 x g for 10 minutes) in a
centrifuge.

6. Store your screwcap tube in the refrigerator until
the next laboratory period (or proceed to step 2
of Lesson 2).

35

Centrifuge

56°C, 10 min

Sheath

Bulb

Water bath

100°C, 5 min

Water bath

Lesson 1 Hair Follicle DNA Template Preparation