User:Karmella Haynes/Notebook/BioBrick cloning/2013/01/09
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* Use gg2 (pSB1A3), gg3, gg4; not gg1 (V0120) since it has BsmBI sites | * Use gg2 (pSB1A3), gg3, gg4; not gg1 (V0120) since it has BsmBI sites | ||
| - | Dilutions | + | * Dilutions |
| - | {| {{table}} border="1" cellspacing="3" <!-- | + | {| {{table}} border="1" cellspacing="3" <!-- Dilution table --> |
|- valign="top" | |- valign="top" | ||
| bgcolor=#cfcfcf | DNA | | bgcolor=#cfcfcf | DNA | ||
| - | | bgcolor=#cfcfcf | length | + | | bgcolor=#cfcfcf | length (bp) |
| - | | bgcolor=#cfcfcf | | + | | bgcolor=#cfcfcf | ng/μL |
| bgcolor=#cfcfcf | Vol. | | bgcolor=#cfcfcf | Vol. | ||
| bgcolor=#cfcfcf | + dH<sub>2</sub>O | | bgcolor=#cfcfcf | + dH<sub>2</sub>O | ||
| Line 32: | Line 32: | ||
|} | |} | ||
| + | |||
| + | * Golden Gate Reactions | ||
| + | # hPCD + BL01 + pSB1A3 | ||
| + | # pSB1A3 | ||
| + | |||
| + | {| {{table}} border="1" cellspacing="3" <!-- Golden Gate Rxn. table --> | ||
| + | |- valign="top" | ||
| + | | Reagent || 1 || 2 | ||
| + | |- | ||
| + | | gg2 pSB1A3 || 1.0 || 1.0 | ||
| + | |- | ||
| + | | gg3 hPCD || 1.0 || --- | ||
| + | |- | ||
| + | | gg4 BL01 || 1.0 || --- | ||
| + | |- | ||
| + | | 10x ligase buffer || 1.0 || 1.0 | ||
| + | |- | ||
| + | | NEB T4 lgase || 0.25 || 0.25 | ||
| + | |- | ||
| + | | BsmBI || 0.5 || 0.5 | ||
| + | |- | ||
| + | | dH<sub>2</sub>O || 5.25 || 7.25 | ||
| + | |- | ||
| + | | || 10.0 || 10.0 | ||
| + | |} | ||
| + | |||
| + | Thermal cycler | ||
| + | * [45°C, 2 min.; 16°C, 5 min.] x25 | ||
| + | * 60°C, 20 min. | ||
| + | * 80°C, 20 min. | ||
Revision as of 21:48, 9 January 2013
 Karmella's BioBrick Cloning
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01/09/13
Golden Gate Assembly
Thermal cycler
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