Team:KIT-Kyoto/Notebook-week6

From 2012.igem.org

(Difference between revisions)
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Line 100: Line 100:
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
<br><br>
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td>pSB1C3-UAS-1, -2, -3, -4</Td><Td>5uL</Td></Tr>
 +
<tr><td>3buffer(NEB)</td><Td>2uL</Td></tr>
 +
<Tr><Td>BglⅡ</Td><Td>0.5uL</Td></Tr>
 +
<td>dH<sub>2</sub>O</td><Td>12.5uL</Td></tr>
 +
<td>Total</td><Td>20uL</Td></tr>
 +
</Table>
 +
<br><BR>
<img src="http://2012.igem.org/wiki/images/0/0c/0911akit.png" width="500" height="300">
<img src="http://2012.igem.org/wiki/images/0/0c/0911akit.png" width="500" height="300">
<br><br>
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td>GAL4</Td><Td>40uL</Td></Tr>
 +
<tr><td>4 buffer(NEB)</td><Td>5uL</Td></tr>
 +
<Tr><Td>XbaⅠ(NEB)</Td><Td>0.5uL</Td></Tr>
 +
<Tr><Td>SpeⅠ(NEB)</Td><Td>0.5uL</Td></Tr><Tr>
 +
<Td>CIP</Td><Td>0.5uL</Td></Tr><Tr>
 +
<Td>100×BSA</Td><Td>0.5uL</Td></Tr>
 +
<td>dH<sub>2</sub>O</td><Td>3uL</Td></tr>
 +
<td>Total</td><Td>40uL</Td></tr>
 +
</Table>
 +
<br><BR>
<img src="http://2012.igem.org/wiki/images/d/da/0911kit.png" width="500" height="300">
<img src="http://2012.igem.org/wiki/images/d/da/0911kit.png" width="500" height="300">
<br><br>
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td></Td><Td>1uL</Td></Tr>
 +
<tr><td></td><Td>2.5uL</Td></tr>
 +
<Tr><Td></Td><Td>2.5uL</Td></Tr>
 +
<td>Ligation high</td><Td>6uL</Td></tr>
 +
<td>Total</td><Td>12uL</Td></tr>
 +
</Table>
 +
<br><BR>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td></Td><Td>2uL</Td></Tr>
 +
<tr><td></td><Td>2.5uL</Td></tr>
 +
<Tr><Td></Td><Td>1.5uL</Td></Tr>
 +
<td>Ligation high</td><Td>6uL</Td></tr>
 +
<td>Total</td><Td>12uL</Td></tr>
 +
</Table>
 +
<br><BR>
<h2>September 12th</h2>
<h2>September 12th</h2>
<br>
<br>
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
<br><br>
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td>pSB1C3-UAS-1, -2</Td><Td>40uL</Td></Tr>
 +
<tr><td>3buffer(NEB)</td><Td>5uL</Td></tr>
 +
<Tr><Td>SpeⅠ</Td><Td>1uL</Td></Tr>
 +
<td>100×BSA</td><Td>0.5uL</Td></tr><tr>
 +
<td>dH<sub>2</sub>O</td><Td>3.5uL</Td></tr>
 +
<td>Total</td><Td>50uL</Td></tr>
 +
</Table>
 +
<br><BR>
<img src="http://2012.igem.org/wiki/images/a/ab/0912kit.png" width="500" height="300">
<img src="http://2012.igem.org/wiki/images/a/ab/0912kit.png" width="500" height="300">
<br><br>
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td></Td><Td>1uL</Td></Tr>
 +
<tr><td></td><Td>2uL</Td></tr>
 +
<Tr><Td>EGFP or LacZ</Td><Td>2uL</Td></Tr>
 +
<td>Ligation high</td><Td>2.5uL</Td></tr><tr>
 +
<td>Total</td><Td>7.5uL</Td></tr>
 +
</Table>
 +
<br><BR>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td></Td><Td>2uL</Td></Tr>
 +
<tr><td></td><Td>2uL</Td></tr>
 +
<Tr><Td></Td><Td>2uL</Td></Tr>
 +
<td>Ligation high</td><Td>6uL</Td></tr><tr>
 +
<td>Total</td><Td>12uL</Td></tr>
 +
</Table>
 +
<br><BR>
<h2>September 13th</h2>
<h2>September 13th</h2>
<br>
<br>
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
<br><br>
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td>DNA sample</Td><Td>1uL</Td></Tr>
 +
<tr><td>4 buffer(NEB)</td><Td>0.5uL</Td></tr>
 +
<Tr><Td>EcoRⅠ-HF(NEB)</Td><Td>0.2uL</Td></Tr>
 +
<Tr><Td>SpeⅠ(NEB)</Td><Td>0.2uL</Td></Tr><Tr>
 +
<Td>100×BSA</Td><Td>0.05uL</Td></Tr>
 +
<td>dH<sub>2</sub>O</td><Td>3.25uL</Td></tr>
 +
<td>Total</td><Td>5uL</Td></tr>
 +
</Table>
 +
<BR><BR>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td>DNA sample</Td><Td>1uL</Td></Tr>
 +
<tr><td>3 buffer(NEB)</td><Td>0.5uL</Td></tr>
 +
<Tr><Td>BglⅡ</Td><Td>0.2uL</Td></Tr>
 +
<td>dH<sub>2</sub>O</td><Td>3.3uL</Td></tr>
 +
<td>Total</td><Td>5uL</Td></tr>
 +
</Table>
<img src="http://2012.igem.org/wiki/images/3/3f/0913kit.png" width="500" height="300">
<img src="http://2012.igem.org/wiki/images/3/3f/0913kit.png" width="500" height="300">
<br><br>
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td></Td><Td>1uL</Td></Tr>
 +
<tr><td></td><Td>1.5uL</Td></tr>
 +
<Tr><Td>dH<sub>2</sub>O</Td><Td>2.5uL</Td></Tr>
 +
<td>Ligation high</td><Td>2.5uL</Td></tr><tr>
 +
<td>Total</td><Td>7.5uL</Td></tr>
 +
</Table>
 +
<br><BR>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td></Td><Td>2.5uL</Td></Tr>
 +
<tr><td></td><Td>2.5uL</Td></tr>
 +
<tr><td></td><Td>2uL</Td></tr>
 +
<td>Ligation high</td><Td>7uL</Td></tr><tr>
 +
<td>Total</td><Td>14uL</Td></tr>
 +
</Table>
 +
<br><BR>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td></Td><Td>1uL</Td></Tr>
 +
<tr><td></td><Td>2uL</Td></tr>
 +
<Tr><Td>dH<sub>2</sub>O</Td><Td>2uL</Td></Tr>
 +
<td>Ligation high</td><Td>5uL</Td></tr><tr>
 +
<td>Total</td><Td>10uL</Td></tr>
 +
</Table>
 +
<br><BR>
<h2>September 14th</h2>
<h2>September 14th</h2>
<br>
<br>
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℃.
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℃.
<br><br>
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td>DNA sample</Td><Td>1uL</Td></Tr>
 +
<tr><td>4 buffer(NEB)</td><Td>1uL</Td></tr>
 +
<Tr><Td>EcoRⅠ-HF(NEB)</Td><Td>0.2uL</Td></Tr>
 +
<Tr><Td>SpeⅠ(NEB)</Td><Td>0.2uL</Td></Tr><Tr>
 +
<Td>100×BSA</Td><Td>0.1uL</Td></Tr>
 +
<td>dH<sub>2</sub>O</td><Td>7.5uL</Td></tr>
 +
<td>Total</td><Td>10uL</Td></tr>
 +
</Table>
<img src="http://2012.igem.org/wiki/images/e/ec/0914kit.png" width="500" height="300">
<img src="http://2012.igem.org/wiki/images/e/ec/0914kit.png" width="500" height="300">
 +
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td></Td><Td>2uL</Td></Tr>
 +
<tr><td></td><Td>3uL</Td></tr>
 +
<tr><td></td><Td>1uL(HS) or 2.5uL(Act5c)</Td></tr>
 +
<td>Ligation high</td><Td>6uL(HS) or 7.5uL(Act5c)</Td></tr><tr>
 +
<td>Total</td><Td>12uL(HS) or 15uL(Act5c)</Td></tr>
 +
</Table>
 +
<br><BR>
<h2>September 15th</h2>
<h2>September 15th</h2>
<br>
<br>
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.
 +
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td>DNA sample</Td><Td>1uL</Td></Tr>
 +
<tr><td>4 buffer(NEB)</td><Td>1uL</Td></tr>
 +
<Tr><Td>EcoRⅠ-HF(NEB)</Td><Td>0.2uL</Td></Tr>
 +
<Tr><Td>SpeⅠ(NEB)</Td><Td>0.2uL</Td></Tr><Tr>
 +
<Td>100×BSA</Td><Td>0.1uL</Td></Tr>
 +
<td>dH<sub>2</sub>O</td><Td>7.5uL</Td></tr>
 +
<td>Total</td><Td>10uL</Td></tr>
 +
</Table>
<br><br>
<br><br>
<img src="http://2012.igem.org/wiki/images/d/d3/0915kit.png" width="500" height="300">
<img src="http://2012.igem.org/wiki/images/d/d3/0915kit.png" width="500" height="300">
Line 135: Line 264:
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.
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.
<br><br>
<br><br>
 +
<Table Border Cellspacing="0">
 +
<Tr><Td>DNA sample</Td><Td>1uL</Td></Tr>
 +
<tr><td>4 buffer(NEB)</td><Td>1uL</Td></tr>
 +
<Tr><Td>EcoRⅠ-HF(NEB)</Td><Td>0.2uL</Td></Tr>
 +
<Tr><Td>SpeⅠ(NEB)</Td><Td>0.2uL</Td></Tr><Tr>
 +
<Td>100×BSA</Td><Td>0.1uL</Td></Tr>
 +
<td>dH<sub>2</sub>O</td><Td>7.5uL</Td></tr>
 +
<td>Total</td><Td>10uL</Td></tr>
 +
</Table>
 +
<br><br>
 +
<img src="http://2012.igem.org/wiki/images/2/26/0917kit.png" width="500" height="300">
</div>
</div>
</div>
</div>

Revision as of 10:11, 23 September 2012






September 11th


The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.

pSB1C3-UAS-1, -2, -3, -45uL
3buffer(NEB)2uL
BglⅡ0.5uL
dH2O12.5uL
Total20uL




GAL440uL
4 buffer(NEB)5uL
XbaⅠ(NEB)0.5uL
SpeⅠ(NEB)0.5uL
CIP0.5uL
100×BSA0.5uL
dH2O3uL
Total40uL




1uL
2.5uL
2.5uL
Ligation high6uL
Total12uL


2uL
2.5uL
1.5uL
Ligation high6uL
Total12uL


September 12th


The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.

pSB1C3-UAS-1, -240uL
3buffer(NEB)5uL
SpeⅠ1uL
100×BSA0.5uL
dH2O3.5uL
Total50uL




1uL
2uL
EGFP or LacZ2uL
Ligation high2.5uL
Total7.5uL


2uL
2uL
2uL
Ligation high6uL
Total12uL


September 13th


The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.

DNA sample1uL
4 buffer(NEB)0.5uL
EcoRⅠ-HF(NEB)0.2uL
SpeⅠ(NEB)0.2uL
100×BSA0.05uL
dH2O3.25uL
Total5uL


DNA sample1uL
3 buffer(NEB)0.5uL
BglⅡ0.2uL
dH2O3.3uL
Total5uL


1uL
1.5uL
dH2O2.5uL
Ligation high2.5uL
Total7.5uL


2.5uL
2.5uL
2uL
Ligation high7uL
Total14uL


1uL
2uL
dH2O2uL
Ligation high5uL
Total10uL


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℃.

DNA sample1uL
4 buffer(NEB)1uL
EcoRⅠ-HF(NEB)0.2uL
SpeⅠ(NEB)0.2uL
100×BSA0.1uL
dH2O7.5uL
Total10uL


2uL
3uL
1uL(HS) or 2.5uL(Act5c)
Ligation high6uL(HS) or 7.5uL(Act5c)
Total12uL(HS) or 15uL(Act5c)


September 15th


The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃.

DNA sample1uL
4 buffer(NEB)1uL
EcoRⅠ-HF(NEB)0.2uL
SpeⅠ(NEB)0.2uL
100×BSA0.1uL
dH2O7.5uL
Total10uL




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.

DNA sample1uL
4 buffer(NEB)1uL
EcoRⅠ-HF(NEB)0.2uL
SpeⅠ(NEB)0.2uL
100×BSA0.1uL
dH2O7.5uL
Total10uL