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Team:UNIPV-Pavia/Notebook/Week5
From 2008.igem.org
(Difference between revisions)
| Line 38: | Line 38: | ||
'''06/16/08''' | '''06/16/08''' | ||
<br> | <br> | ||
| - | *We picked up the only colony in '''BBa_B0030'''-BBa_C0078 plate and | + | *We picked up the only colony in '''BBa_B0030'''-BBa_C0078 plate to grow a 9 ml culture of transformed bacteria overnight. |
| + | |||
| + | *We also infected 9 ml of LB + suitable antibiotic with 30 µl of: | ||
| + | {|cellpadding="20px" | ||
| + | |BBa_B0030 | ||
| + | |BBa_E1010 | ||
| + | |BBa_E0040 | ||
| + | |- | ||
| + | |BBa_C0061 | ||
| + | |BBa_C0051 | ||
| + | |} | ||
| + | *glycerol stocks. We incubated the 9 ml culture overnight at 37°C, 220 rpm. | ||
| + | |||
| + | <br><br> | ||
| + | '''06/17/08''' | ||
| + | <br> | ||
| + | *Glycerol stocks for: | ||
| + | {|cellpadding="20px" | ||
| + | |BBa_B0030 | ||
| + | |BBa_E1010 | ||
| + | |BBa_E0040 | ||
| + | |- | ||
| + | |BBa_C0061 | ||
| + | |BBa_C0051 | ||
| + | |'''BBa_B0030'''-BBa_C0078 | ||
| + | |} | ||
| + | |||
| + | *Miniprep for all these parts. | ||
| + | |||
| + | *Digestion for: | ||
| + | {|cellpadding="20px" | ||
| + | |BBa_B0030 (S-P) | ||
| + | |BBa_E1010 (X-P) | ||
| + | |BBa_E0040 (X-P) | ||
| + | |- | ||
| + | |BBa_C0061 (X-P) | ||
| + | |BBa_C0051 (X-P) | ||
| + | |'''BBa_B0030'''-BBa_C0078 (X-S) | ||
| + | |} | ||
| + | |||
| + | *Gel run for '''BBa_B0030'''-BBa_C0078 to check for insert length: unfortunately, there was not a band where we expected...the only colony was a false positive. We'll try to ligate it in the next days. | ||
| + | |||
| + | *Gel run for: | ||
| + | {|cellpadding="20px" | ||
| + | |BBa_B0030 (S-P) | ||
| + | |BBa_E1010 (X-P) | ||
| + | |BBa_E0040 (X-P) | ||
| + | |- | ||
| + | |BBa_C0061 (X-P) | ||
| + | |BBa_C0051 (X-P) | ||
| + | |} | ||
| + | |||
| + | *Gel cut and DNA extraction. | ||
| + | |||
| + | *We put DNA at -20°C. The next day we will perform some ligation reaction in different conditions, looking for the best protocol. | ||
| + | |||
| + | <br><br> | ||
| + | '''06/18/08''' | ||
| + | <br> | ||
| + | *We gave a lecture about Synthetic Biology and our current work at DIS (Department of Informatics and System Science). | ||
| + | |||
| + | *Antarctic Phosphatase for half of BBa_B0030 (S-P) volume. | ||
| + | |||
| + | *We planned the following ligation experiments: | ||
| + | **Transformation with BBa_B0030(S-P), to check background noise (we will know the amount of not digested vector); | ||
| + | **Transformation with this ligation: BBa_B0030(S-P) after Antarctic Phosphatase treatment and no insert; | ||
| + | **Transformation with this ligation: BBa_B0030(S-P) without Antarctic Phosphatase treatment and no insert; | ||
| + | **Transformation with this ligation: BBa_B0030(S-P) without Antarctic Phosphatase treatment and BBa_C0061 insert; | ||
| + | **Transformation with this ligation: BBa_B0030(S-P) after Antarctic Phosphatase treatment and BBa_C0061 insert; | ||
| + | **3 Transformation with these ligations: BBa_B0030(S-P) after Antarctic Phosphatase treatment and BBa_C0051, BBa_E1010, BBa_E0040 inserts; | ||
| + | |||
| + | *Antarctic Phosphatase for half of BBa_B0030 (S-P) volume. | ||
| + | |||
| + | *We transformed 60 µl of TOP10 with 1 µl of BBa_B0030 (S-P) | ||
| + | |||
| + | *We plated transformed bacteria and incubated them at 37°C overnight. | ||
| + | |||
| + | *Antarctic Phosphatase for half of BBa_B0030 (S-P) volume. | ||
| + | |||
| + | *Ligation: | ||
Revision as of 16:25, 6 July 2008
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|---|---|---|---|---|
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Notebook
| Week 1 | Week 2 | Week 3 | Week 4 | Week 5 | Week 6 | Week 7 |
|---|
Week 5: 06/16/08 - 06/20/08
06/16/08
- We picked up the only colony in BBa_B0030-BBa_C0078 plate to grow a 9 ml culture of transformed bacteria overnight.
- We also infected 9 ml of LB + suitable antibiotic with 30 µl of:
| BBa_B0030 | BBa_E1010 | BBa_E0040 |
| BBa_C0061 | BBa_C0051 |
- glycerol stocks. We incubated the 9 ml culture overnight at 37°C, 220 rpm.
06/17/08
- Glycerol stocks for:
| BBa_B0030 | BBa_E1010 | BBa_E0040 |
| BBa_C0061 | BBa_C0051 | BBa_B0030-BBa_C0078 |
- Miniprep for all these parts.
- Digestion for:
| BBa_B0030 (S-P) | BBa_E1010 (X-P) | BBa_E0040 (X-P) |
| BBa_C0061 (X-P) | BBa_C0051 (X-P) | BBa_B0030-BBa_C0078 (X-S) |
- Gel run for BBa_B0030-BBa_C0078 to check for insert length: unfortunately, there was not a band where we expected...the only colony was a false positive. We'll try to ligate it in the next days.
- Gel run for:
| BBa_B0030 (S-P) | BBa_E1010 (X-P) | BBa_E0040 (X-P) |
| BBa_C0061 (X-P) | BBa_C0051 (X-P) |
- Gel cut and DNA extraction.
- We put DNA at -20°C. The next day we will perform some ligation reaction in different conditions, looking for the best protocol.
06/18/08
- We gave a lecture about Synthetic Biology and our current work at DIS (Department of Informatics and System Science).
- Antarctic Phosphatase for half of BBa_B0030 (S-P) volume.
- We planned the following ligation experiments:
- Transformation with BBa_B0030(S-P), to check background noise (we will know the amount of not digested vector);
- Transformation with this ligation: BBa_B0030(S-P) after Antarctic Phosphatase treatment and no insert;
- Transformation with this ligation: BBa_B0030(S-P) without Antarctic Phosphatase treatment and no insert;
- Transformation with this ligation: BBa_B0030(S-P) without Antarctic Phosphatase treatment and BBa_C0061 insert;
- Transformation with this ligation: BBa_B0030(S-P) after Antarctic Phosphatase treatment and BBa_C0061 insert;
- 3 Transformation with these ligations: BBa_B0030(S-P) after Antarctic Phosphatase treatment and BBa_C0051, BBa_E1010, BBa_E0040 inserts;
- Antarctic Phosphatase for half of BBa_B0030 (S-P) volume.
- We transformed 60 µl of TOP10 with 1 µl of BBa_B0030 (S-P)
- We plated transformed bacteria and incubated them at 37°C overnight.
- Antarctic Phosphatase for half of BBa_B0030 (S-P) volume.
- Ligation:
