Team:KIT-Kyoto/Notebook-week5
From 2012.igem.org
(Difference between revisions)
Line 106: | Line 106: | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> 9/3 GAL4 </Td><Td> 40uL </Td></Tr> |
- | <tr><td> | + | <tr><td> M buffer(TOYOBO) </td><td> 5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> XbaⅠ(TOYOBO) </Td><Td> 0.5uL </Td></Tr> |
- | <tr><td> | + | <tr><td> SpeⅠ(TOYOBO) </td><td> 0.5uL </td></tr><tr> |
- | <td> | + | <td> dH<sub>2</sub>O </td><td> 4uL </td></tr><tr> |
- | <td> | + | <td> Total </td><td> 50uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> each DNA </Td><Td> 40uL </Td></Tr> |
- | <tr><td> | + | <tr><td> 3 buffer(NEB) </td><td> 5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> BglⅡ(NEB) </Td><Td> 0.5uL </Td></Tr><tr> |
- | <td> | + | <td> dH<sub>2</sub>O </td><td> 4.5uL </td></tr><tr> |
- | <td> | + | <td> Total </td><td> 50uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> 9/3 GAL4 </Td><Td> 23uL </Td></Tr> |
- | <tr><td> | + | <tr><td> M buffer(TOYOBO) </td><td> 10uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> XbaⅠ(TOYOBO) </Td><Td> 1uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> SpeⅠ(TOYOBO) </Td><Td> 1uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> CIP </Td><Td> 0.5uL </Td></Tr> |
- | <td> | + | <td> dH<sub>2</sub>O </td><td> 64.5uL </td></tr> |
- | <td> | + | <td> Total </td><td> 100uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/b/b0/0904kit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/b/b0/0904kit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
Line 145: | Line 147: | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> EGFP </Td><Td> pSB1C3 </Td></Tr> |
- | <tr><td> | + | <tr><td> DNA template </td><td> 40uL </td><Td> 23uL </Td></tr> |
- | <Tr><Td> | + | <Tr><Td> 3 buffer </Td><Td> 5uL< /Td><Td> 5uL </Td></Tr> |
- | <tr><td> | + | <tr><td> BglⅡ </td><td> 0.5uL </td><Td> 0.5uL </Td></tr><tr> |
- | <td> | + | <td> dH<sub>2</sub>O </td><td> 4.5uL </td><Td> 21.5uL </Td></tr> |
- | <td> | + | <td> Total </td><td> 50uL </td><Td> 50uL </Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/3/3a/0905akit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/3/3a/0905akit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> DNA template </Td><Td> 40uL </Td></Tr> |
- | <tr><td> | + | <tr><td> 4 buffer(NEB) </td><td> 5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> SpeⅠ </Td><Td> 0.5uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 100×BSA </Td><Td> 0.5uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> CIP </Td><Td> 0.5uL </Td></Tr> |
- | <td> | + | <td> dH<sub>2</sub>O </td><td> 4uL </td></tr> |
- | <td> | + | <td> Total </td><td> 50uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> 1uL </Td></Tr> |
- | <tr><td></td><td> | + | <tr><td></td><td> 1uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> dH<sub>2</sub>O </Td><Td> 3uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> Ligation high </Td><Td> 2.5uL </Td></Tr> |
- | <td> | + | <td> Total </td><td> 7.5uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
Line 188: | Line 192: | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> All samples </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> DNA template </Td><Td> 0.25uL </Td></Tr> |
- | <tr><td> | + | <tr><td> 10× KOD plus buffer </td><td> 5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> 2mM dNTPs </Td><Td> 5uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr><tr> |
- | <td> | + | <td> 10P 5’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> KOD plus </td><td> 1uL </td></tr> |
- | <Tr><td> | + | <Tr><td> dH<sub>2</sub>O </td><td> 34.15uL </td></tr> |
- | <Tr><td> | + | <Tr><td> Total </td><td> 50uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> |
- | <Tr><Td> | + | <Tr><Td> 95℃ </Td><Td> 15sec</Td><Td> 30 cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 58℃ </td><td> 30sec</td><Td> 30 cycle </Td></tr><tr> |
- | <td> | + | <td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td> 30 cycle </Td></tr> |
- | <tr><td> | + | <tr><td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td></Td></tr> |
- | <tr><td> | + | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/5/53/0906kit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/5/53/0906kit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> pSB1C3(XbaⅠ and SpeⅠ cut) </Td><Td> 1uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> GAL4(XbaⅠ and SpeⅠ cut) </Td><Td> 1uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> dH<sub>2</sub>O </td><td> 3uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> Ligation high </Td><Td> 2.5uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> Total </td><td> 7.5uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
Line 233: | Line 239: | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> All samples </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> DNA template </Td><Td> 0.25uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> 2mM dNTPs </td><td> 5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> 10P 5’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> KOD plus </td><td> 1uL </td></tr> |
- | <Tr><td> | + | <Tr><td> dH<sub>2</sub>O </td><td> 34.15uL </td></tr> |
- | <Tr><td> | + | <Tr><td> Total </td><td> 50uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> |
- | <Tr><Td> | + | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 58℃ </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> |
- | <td> | + | <td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td> 30 cycle </Td></tr> |
- | <tr><td> | + | <tr><td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td></Td></tr> |
- | <tr><td> | + | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/4/4c/0907akit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/4/4c/0907akit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
Line 272: | Line 280: | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> All samples </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> DNA template </Td><Td> 1uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> 2mM dNTPs </td><td> 5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> 10P 5’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> KOD plus </td><td> 1uL </td></tr> |
- | <Tr><td> | + | <Tr><td> dH<sub>2</sub>O </td><td> 33.4uL </td></tr> |
- | <Tr><td> | + | <Tr><td> Total </td><td> 50uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> Temperature</Td><Td> Time </Td><Td> Cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> |
- | <Tr><Td> | + | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 58℃ </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> |
- | <td> | + | <td> 68℃ </td></td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td> 30 cycle </Td></tr> |
- | <tr><td> | + | <tr><td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td></Td></tr> |
- | <tr><td> | + | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
Line 299: | Line 308: | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> All samples </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> DNA template </Td><Td> 1uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> 2mM dNTPs </td><td> 5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> 10P 5’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> KOD plus </td><td> 1uL </td></tr> |
- | <Tr><td> | + | <Tr><td> dH<sub>2</sub>O </td><td> 33.4uL </td></tr> |
- | <Tr><td> | + | <Tr><td> Total </td><td> 50uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> |
- | <Tr><Td> | + | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 55℃(-1) or 58℃(-2) </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> |
- | <td> | + | <td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td> 30 cycle </Td></tr> |
- | <tr><td> | + | <tr><td> 68℃ </td><td> 30sec(UAS) or 2min10sec(Except for UAS) </td><Td></Td></tr> |
- | <tr><td> | + | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
Line 336: | Line 346: | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> UAS </Td><Td> 40uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 4 buffer(NEB) </Td><Td> 5uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> EcoRⅠ-HF(NEB) </Td><Td> 0.5uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> SpeⅠ(NEB </td><td> 0.5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> 100×BSA </Td><Td> 0.5uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> dH<sub>2</sub>O </td><td> 3.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> Total </td><td> 50uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/9/91/0909bkit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/9/91/0909bkit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> All sample </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> DNA template </Td><Td> 1uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> 2mM dNTPs <td> 5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> 25mM MgSO4 </Td><Td> 1.6uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> 10P 5’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> 10P 3’ primer </td><td> 1.5uL </td></tr> |
- | <Tr><td> | + | <Tr><td> KOD plus </td><td> 1uL </td></tr> |
- | <Tr><td> | + | <Tr><td> dH<sub>2</sub>O </td><td> 33.4uL </td></tr> |
- | <Tr><td> | + | <Tr><td> Total </td><td> 50uL </td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> |
- | <Tr><Td> | + | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 58℃ </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> |
- | <td> | + | <td> 68℃ </td><td> 2min10sec </td><Td> 30 cycle </Td></tr> |
- | <tr><td> | + | <tr><td> 68℃ </td><td> 2min10sec </td><Td></Td></tr> |
- | <tr><td> | + | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/b/bb/0909ckit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/b/bb/0909ckit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> 1uL </Td></Tr> |
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> 1uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> dH<sub>2</sub>O </Td><Td> 3uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> Ligation high </td><td> 2.5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> Total </Td><Td> 7.5uL </Td></Tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> 1uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> dH<sub>2</sub>O </Td><Td> 4uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> Ligation high </td><td> 2.5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> Total </Td><Td> 7.5uL </Td></Tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
+ | |||
<h2>September 10th</h2> | <h2>September 10th</h2> | ||
<br> | <br> | ||
Line 398: | Line 415: | ||
<br><br> | <br><br> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> HS or Act5c </Td><Td> 40uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 4 buffer(NEB) </Td><Td> 5uL </Td></Tr> |
- | <Tr><td> | + | <Tr><td> XbaⅠ-HF(NEB) <td> 0.5uL </td></tr> |
- | <Tr><Td> | + | <Tr><Td> BamHⅠ(NEB )</Td><Td> 0.5uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 100×BSA </Td><Td> 0.5uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> dH<sub>2</sub>O </Td><Td> 3.5uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> Total </Td><Td> 50uL </Td></Tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/2/27/0910akit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/2/27/0910akit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> G-1 and G-2 </Td><Td> G-3 and G-4 </Td></Tr> |
- | <tr><td> | + | <tr><td> DNA template </td><td> 1uL </td><Td> 4uL </Td></tr> |
- | <Tr><Td> | + | <Tr><Td> 10× KOD plus buffer </Td><Td> 5uL </Td><Td> 5uL </Td></Tr> |
- | <tr><td> | + | <tr><td> 2mM dNTPs </td><td> 5uL </td><Td> 5uL </Td></tr><tr> |
- | <td> | + | <td> 25mM MgSO4 </td><td> 1.6uL </td><Td> 1.6uL </Td></tr> |
- | <tr><td> | + | <tr><td> 10P 5’ primer </td><td> 1uL </td><Td> 1uL </Td></tr> |
- | <tr><td> | + | <tr><td> 10P 3’ primer </td><td> 1uL </td><Td> 1uL </Td></tr> |
- | <tr><td> | + | <tr><td> KOD plus </td><td> 1uL </td><Td> 1uL </Td></tr> |
- | <tr><td> | + | <tr><td> dH<sub>2</sub>O </td><td> 33.4uL </td><Td> 31.4uL </Td></tr> |
- | <tr><td> | + | <tr><td> Total </td><td> 50uL </td><Td> 50uL </Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> Temperature </Td><Td> Time </Td><Td> Cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 95℃ </td><td> 2min </td><Td></Td></tr> |
- | <Tr><Td> | + | <Tr><Td> 95℃ </Td><Td> 15sec </Td><Td> 30 cycle </Td></Tr> |
- | <tr><td> | + | <tr><td> 57℃(G-1 and G-3) or 59℃(G-2 and G-4) </td><td> 2min30sec </td><Td> 30 cycle </Td></tr><tr> |
- | <td> | + | <td> 68℃ </td><td> 2min10sec </td><Td> 30 cycle </Td></tr> |
- | <tr><td> | + | <tr><td> 68℃ </td><td> 2min10sec </td><Td></Td></tr> |
- | <tr><td> | + | <tr><td> 14℃ </td><td> ∞ </td><Td></Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/e/eb/0910bkit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/e/eb/0910bkit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> pSB1C3(PCR) </Td><Td> 40uL </Td></Tr> |
- | <tr><td> | + | <tr><td> 4 buffer(NEB) </td><Td> 5uL </Td></tr> |
- | <Tr><Td> | + | <Tr><Td> EcoRⅠ-HF(NEB) </Td><Td> 0.5uL </Td></Tr> |
- | <tr><td> | + | <tr><td> SpeⅠ(NEB) </td><Td> 0.5uL </Td></tr><tr> |
- | <td> | + | <td> 2100×BSA </td><Td> 0.5uL </Td></tr> |
- | <tr><td> | + | <tr><td> dH<sub>2</sub>O <Td> 3.5uL </Td></tr> |
- | <tr><td> | + | <tr><td> Total </td><Td> 50uL </Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> 13uL </Td></Tr> |
- | <tr><td> | + | <tr><td> 3 buffer(NEB) </td><Td> 2uL </Td></tr> |
- | <Tr><Td> | + | <Tr><Td> BglⅡ(NEB) </Td><Td> 0.5uL </Td></Tr> |
- | <tr><td> | + | <tr><td> dH<sub>2</sub>O <Td> 2.5uL </Td></tr> |
- | <tr><td> | + | <tr><td> Total </td><Td> 20uL </Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/8/86/0910ckit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/8/86/0910ckit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> GAL4 </Td><Td> 20uL </Td></Tr> |
- | <tr><td> | + | <tr><td> 4 buffer(NEB) </td><Td> 4uL </Td></tr> |
- | <Tr><Td> | + | <Tr><Td> XbaⅠ(NEB) </Td><Td> 0.7uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> SpeⅠ(NEB) </Td><Td> 0.7uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 100×BSA </Td><Td> 0.4uL </Td></Tr> |
- | <tr><td> | + | <tr><td> dH<sub>2</sub>O </td><Td> 14.6uL </Td></tr> |
- | <tr><td> | + | <tr><td> Total </td><Td> 40uL </Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/3/39/0910dkit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/3/39/0910dkit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> GAL4 </Td><Td> 40uL </Td></Tr> |
- | <tr><td> | + | <tr><td> 3 buffer(NEB) </td><Td> 5uL </Td></tr> |
- | <Tr><Td> | + | <Tr><Td> BglⅡ(NEB) </Td><Td> 0.5uL </Td></Tr> |
- | <tr><td> | + | <tr><td> dH<sub>2</sub>O </td><Td> 4.5uL </Td></tr> |
- | <tr><td> | + | <tr><td> Total </td><Td> 50uL </Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td> | + | <Tr><Td> GAL4(Bgl Ⅱ cut) </Td><Td> 40uL </Td></Tr> |
- | <tr><td> | + | <tr><td> 4 buffer(NEB) </td><Td> 5uL </Td></tr> |
- | <Tr><Td> | + | <Tr><Td> SpeⅠ </Td><Td> 0.5uL </Td></Tr> |
- | <Tr><Td> | + | <Tr><Td> 100×BSA </Td><Td> 0.5uL </Td></Tr> |
- | <tr><td> | + | <tr><td> dH<sub>2</sub>O </td><Td> 4uL </Td></tr> |
- | <tr><td> | + | <tr><td> Total </td><Td> 50uL </Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<img src="https://static.igem.org/mediawiki/2012/6/6d/0910ekit.png" width="500" height="300"> | <img src="https://static.igem.org/mediawiki/2012/6/6d/0910ekit.png" width="500" height="300"> | ||
<br><br> | <br><br> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> 1uL </Td></Tr> |
- | <tr><td></td><Td> | + | <tr><td></td><Td> 2uL </Td></tr> |
- | <Tr><Td> | + | <Tr><Td> EGFP or LacZ </Td><Td> 2uL </Td></Tr> |
- | <tr><td> | + | <tr><td> Ligation high </td><Td> 5uL </Td></tr> |
- | <tr><td> | + | <tr><td> Total </td><Td> 10uL </Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> | ||
+ | |||
<Table Border Cellspacing="0"> | <Table Border Cellspacing="0"> | ||
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> 1uL </Td></Tr> |
- | <tr><td></td><Td> | + | <tr><td></td><Td> 2uL </Td></tr> |
- | <Tr><Td></Td><Td> | + | <Tr><Td></Td><Td> 2uL </Td></Tr> |
- | <tr><td> | + | <tr><td> Ligation high </td><Td> 5uL </Td></tr> |
- | <tr><td> | + | <tr><td> Total </td><Td> 10uL </Td></tr> |
</Table> | </Table> | ||
<br><BR> | <br><BR> |
Revision as of 02:29, 24 September 2012
September 4thTNFA and API2 The female-virgin flies (yw) and male flies (yw) were collected for mating with the microinjected males and females. Parts
September 5thTNFA and API2 The female-virgin flies (yw) and male flies (yw) were collected and separately kept at 25℃ for mating with the microinjected males and females. Parts
September 6thTNFA and API2 The female-virgin flies (yw) and male flies (yw) were collected and separately kept at 25℃ for mating with the microinjected males and females. Parts
September 7thTNFA and API2 The eclosed male and virgin female flies from microinjected embryos were collected and kept at 25℃ for maturation. Parts
September 8thTNFA and API2 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 eclosed male and virgin female flies from microinjected embryos were again collected and kept at 25℃ for maturation. The female-virgin flies (yw) and male flies (yw) were collected and separately kept at 25℃. Parts
September 9thTNFA and API2 The eclosed male and virgin female flies from microinjected embryos were again collected and kept at 25℃ for maturation. In total 83 flies were eclosed from the microinjected embryos. The calculated viability for the microinjected flies was 12.0%. The female-virgin flies (yw) and male flies (yw) were again collected and separately kept at 25℃. Parts
September 10thTNFA and API2 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. Parts
|