|
Timetable for the workshop
on "Staining of root tip cells in Bawang
Merah (Allium ascalonicu)"
by the Fuelgen Staining Method |
|
|
Time |
Procedure |
Duration |
Material / Equipments |
|
8.30 |
Explanation of the Feulgen Staining Method |
60 min |
|
|
09.30 |
Preparation of chemical solution used here |
20 min |
|
|
09.50 |
Fixation of root tips with ethyl
alcohol:acetic acide(3:1) |
10 min |
(Do not use
these roots for staining today) |
|
10.00 |
Snack |
30 min |
|
|
10.30 |
Root tips are stained by the Feulgen
staining method as mentioned below: |
| |
|
1 |
Root tips in the bottle are washed 5
times with tap water by changing water |
20 min |
Pipette |
|
10.50 |
2 |
About 1 ml of 1N HCl is added and
the bottle is placed on the bench for 3 min |
05 min |
1N HCl |
|
10.55 |
3 |
The bottle is removed to a water bath
and root tips are hydrolyze for 8 min at 60°C |
08 min |
Water bath (set at 60°C) |
|
11.05 |
4 |
Root tips
in the bottle are washed 5 times with tap water by changing water |
20 min |
Pipette |
|
11.25 |
5 |
About 1ml of Schiff's reagent is added
and the bottle is kept in the dark for at least 1 hour |
60 min
|
Schiff's Reagent, Pipette |
|
11.30 |
Lunch Break |
60 min |
|
|
12.30 |
6 |
Root tips are washed 3
times with tap water by changing water |
10 min |
Pipette |
|
12.35 |
7 |
About 1 ml of 45% acetic
acid solution is added |
05 min |
45% Acetic acid |
|
|
A
root tip is squashed as showed below |
05
min/slide |
Microscope slide, cover
slip, Tissue paper, Forceps, Cutter |
|
|
1 |
Drop 45%
acetic acid solution on a microscope slide and put a
root tip on it |
|
|
| 2 |
Cut root from
the tip about 2 mm and lightly stained portion is
removed from the slide |
|
|
| 3 |
Drop 45%
acetic acid solution and put a cover slip on it |
|
|
| 4 |
Spread cells
on the slide by tapping cover slip gently with a finger |
|
|
| 5 |
Put the slide
between tissue papers and squash it with a finger |
|
|
| 6 |
Put the slide
on the microscope stage and bring it into focus |
|
|
| 7 |
Check the
slide if the cells pile up or not |
|
|
|
12.45 |
Squashed slides are placed in a freezer at -20°C
for 30 minutes and cover slips are
removed carefully by using a cutter. |
40 min |
-20°C
Freezer, Cutter |
|
13.25 |
The slides are dried in air |
10 min |
|
|
13.35 |
The slides are dehydrated by dipping them into 25, 50,
70, 85, and 99.5% ethyl alcohol for 10 minutes each. |
1 h |
25, 50, 70, 85 and 99.5%
ethyl alcohol, Staining jar or Beaker, Forceps |
|
14.35
|
Slides are treated with xylen for 10 minutes, mounted
with Canada balsam and covered with cover slips |
10 min |
Xylen, Staining jar or
Beaker, Canada balsam, cover slip |
|
14.45 |
Wash glassware and clean the bench |
10 min |
|
|
14.55 |
Closing |
