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Team:Cambridge-JIC/Guide/Materials/Media
From 2014.igem.org
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| - | + | =Media= | |
| + | ==Agrobacterium== | ||
| + | For plates, | ||
Use standard LB + 1.5% agar + required antibiotics. | Use standard LB + 1.5% agar + required antibiotics. | ||
| - | + | ||
| + | ==Marchantia== | ||
| + | ===Marchantia plates=== | ||
For marchantia, 1/2 Gamborg's B5 media + 1.2% agar is effective. Gamborg's B5 salts can be found readily from chemical suppliers. | For marchantia, 1/2 Gamborg's B5 media + 1.2% agar is effective. Gamborg's B5 salts can be found readily from chemical suppliers. | ||
| Line 9: | Line 13: | ||
Always seal all plates using micropore tape. | Always seal all plates using micropore tape. | ||
| - | + | ===Selection plates=== | |
| - | If using the Marchantia polymorpha resistance cassette and | + | If using the Marchantia polymorpha resistance cassette and Agrobacterium mediated transformation, use Marchantia plates with 10ug/ml hygromycin and 100ug/ml cefotaxime (to ensure that agrobacterium does not colonize the plate). |
Note: selection plates do not contain a high enough concentration of hygromycin to preclude fungal colonisation. Work in sterile conditions. | Note: selection plates do not contain a high enough concentration of hygromycin to preclude fungal colonisation. Work in sterile conditions. | ||
| - | + | ==Miscellaneous== | |
| + | ===Acetosyringone=== | ||
Use a fresh stock, with a concentration of 100mM, in DMSO. This will be used in transformation protocols as 1000x. | Use a fresh stock, with a concentration of 100mM, in DMSO. This will be used in transformation protocols as 1000x. | ||
Revision as of 00:39, 17 October 2014
Contents |
Media
Agrobacterium
For plates, Use standard LB + 1.5% agar + required antibiotics.
Marchantia
Marchantia plates
For marchantia, 1/2 Gamborg's B5 media + 1.2% agar is effective. Gamborg's B5 salts can be found readily from chemical suppliers.
Always use freshly autoclaved media, and work in a laminar flow hood, maintaining sterile conditions.
Fungal spores rapidly colonise the plates under conditions for Marchantia growth.
Always seal all plates using micropore tape.
Selection plates
If using the Marchantia polymorpha resistance cassette and Agrobacterium mediated transformation, use Marchantia plates with 10ug/ml hygromycin and 100ug/ml cefotaxime (to ensure that agrobacterium does not colonize the plate).
Note: selection plates do not contain a high enough concentration of hygromycin to preclude fungal colonisation. Work in sterile conditions.
Miscellaneous
Acetosyringone
Use a fresh stock, with a concentration of 100mM, in DMSO. This will be used in transformation protocols as 1000x.
