Haynes Lab:Notebook/Engineering PC-TFs/2013/01/25
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| - | == | + | ==01/25/13== |
| - | * | + | * Type IIS assembly - day 1 |
| + | |||
| + | |||
| + | ---- | ||
| + | |||
| + | PCR | ||
| + | * Resuspend new primers to a final concentration of 100 μM | ||
| + | * Make 10 μM dilutions in separate tubes | ||
| + | * All of the reactions are as follows: | ||
| + | |||
| + | # template, primer 1, primer 2 | ||
| + | # template, primer 1, primer 2 | ||
| + | # template, primer 1, primer 2 | ||
| + | # template, primer 1, primer 2 | ||
| + | |||
| + | |||
| + | {| {{table}} border="1" cellspacing="3" <!-- PCR rxn. table --> | ||
| + | |- valign="top" | ||
| + | | bgcolor=#cfcfcf | Reagent | ||
| + | | bgcolor=#cfcfcf | Volume | ||
| + | | rowspan="7" | <u>Expected:</u><br>1. pSB1A3 = ~2000<br>2. hPCD = 186<br>3. BL01 = 2520 | ||
| + | | rowspan="7" | [[Image:KAH011013_gel1.jpg|200px|Hover name]]<br>10 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder] | ||
| + | |- | ||
| + | | DNA(plasmid) || 0.2 μL | ||
| + | |- | ||
| + | | primer 1 || 1.0 | ||
| + | |- | ||
| + | | primer 2 || 1.0 | ||
| + | |- | ||
| + | | 2x GoTaq || 25.0 | ||
| + | |- | ||
| + | | dH<sub>2</sub>O || 22.8 | ||
| + | |- | ||
| + | | || 50.0 μL | ||
| + | |} | ||
| + | |||
| + | PCR | ||
| + | * 95°C, 3 min. | ||
| + | * [95°C, 30 sec.; 57°C, 30 sec.; 72°C, 1 min.] x35 | ||
| + | * 72°C, 3 min. | ||
| + | * 4°C, ∞ | ||
| + | |||
Revision as of 16:38, 25 January 2013
 Engineering PC-TFs
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