September 11th
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
Parts
pSB1C3-UAS-1, -2, -3, -4 | 5uL |
3buffer(NEB) | 2uL |
BglⅡ | 0.5uL |
dH2O | 12.5uL |
Total | 20uL |
GAL4 | 40uL |
4 buffer(NEB) | 5uL |
XbaⅠ(NEB) | 0.5uL |
SpeⅠ(NEB) | 0.5uL |
CIP | 0.5uL |
100×BSA | 0.5uL |
dH2O | 3uL |
Total | 40uL |
| 1uL |
| 2.5uL |
| 2.5uL |
Ligation high | 6uL |
Total | 12uL |
| 2uL |
| 2.5uL |
| 1.5uL |
Ligation high | 6uL |
Total | 12uL |
September 12th
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
Parts
pSB1C3-UAS-1, -2 | 40uL |
3buffer(NEB) | 5uL |
SpeⅠ | 1uL |
100×BSA | 0.5uL |
dH2O | 3.5uL |
Total | 50uL |
| 1uL |
| 2uL |
EGFP or LacZ | 2uL |
Ligation high | 2.5uL |
Total | 7.5uL |
| 2uL |
| 2uL |
| 2uL |
Ligation high | 6uL |
Total | 12uL |
September 13th
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
Parts
DNA sample | 1uL |
4 buffer(NEB) | 0.5uL |
EcoRⅠ-HF(NEB) | 0.2uL |
SpeⅠ(NEB) | 0.2uL |
100×BSA | 0.05uL |
dH2O | 3.25uL |
Total | 5uL |
DNA sample | 1uL |
3 buffer(NEB) | 0.5uL |
BglⅡ | 0.2uL |
dH2O | 3.3uL |
Total | 5uL |
| 1uL |
| 1.5uL |
dH2O | 2.5uL |
Ligation high | 2.5uL |
Total | 7.5uL |
| 2.5uL |
| 2.5uL |
| 2uL |
Ligation high | 7uL |
Total | 14uL |
| 1uL |
| 2uL |
dH2O | 2uL |
Ligation high | 5uL |
Total | 10uL |
September 14th
The adult male and virgin female flies from microinjected embryos were mated with yw virgin female flies and yw male flies, respectively. Mated flies were transferred to the new food vials in every three days to lay eggs as much as possible. The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
Parts
DNA sample | 1uL |
4 buffer(NEB) | 1uL |
EcoRⅠ-HF(NEB) | 0.2uL |
SpeⅠ(NEB) | 0.2uL |
100×BSA | 0.1uL |
dH2O | 7.5uL |
Total | 10uL |
| 2uL |
| 3uL |
| 1uL(HS) or 2.5uL(Act5c) |
Ligation high | 6uL(HS) or 7.5uL(Act5c) |
Total | 12uL(HS) or 15uL(Act5c) |
September 15th
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
Parts
DNA sample | 1uL |
4 buffer(NEB) | 1uL |
EcoRⅠ-HF(NEB) | 0.2uL |
SpeⅠ(NEB) | 0.2uL |
100×BSA | 0.1uL |
dH2O | 7.5uL |
Total | 10uL |
September 16th
The adult male and virgin female flies from microinjected embryos were mated with yw virgin female flies and yw male flies, respectively. In total 83 flies from microinjected embryos were mated with yw flies. Mated flies were transferred to the new food vials in every three days to lay eggs as much as possible.
September 17th
The progeny flies were inspected by dissecting microscope to look for the successfully transformed w+ red eye flies (red eye screening). However, no red eye fly was found.
Parts
DNA sample | 1uL |
4 buffer(NEB) | 1uL |
EcoRⅠ-HF(NEB) | 0.2uL |
SpeⅠ(NEB) | 0.2uL |
100×BSA | 0.1uL |
dH2O | 7.5uL |
Total | 10uL |