User:Behzad Damadzadeh/Notebook/PcTF Subcloning in E-coli/2013/10/10: Difference between revisions
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* Run 5 μL of each PCR reaction on 10% agarose gel. | * Run 5 μL of each PCR reaction on 10% agarose gel. | ||
[[Image:BD002-SD-SAPI.png |frame|none|alt=Alt text|Columns 1,3 amplified show BD002 ~6300 bp ]]. | [[Image:BD002-SD-SAPI.png |frame|none|alt=Alt text|Columns 1,3 amplified show BD002 ~6300 bp.]] | ||
* Using the GenElute™ PCR Clean-Up Kit for rapid purification of double-stranded PCR amplification products. | |||
** The purified result: Column1: 100ng/μL and Column 3: 28ng/μL. | |||
* Cutting the SAPI sites to make the sticky ends for ligation: | |||
{| {{table}} cellspacing="3" width=350px | |||
| Amplified DNA (500ng) || 5.00 μl || 17.00 μL | |||
|- | |||
| SAPI || 1.0 μl || 1.0 μL | |||
|- | |||
| SpeI || 1.0 μl | |||
|- | |||
| 10x buffer || 5.00 μl || 5.00 μL | |||
|- | |||
| dH<sub>2</sub>O || 39.00 μl || 27.00 μl | |||
|- | |||
| Total || 50.0 μl || 50.0 μl | |||
|- | |||
| Incubate at 37°C overnight. | |||
|} | |||
* Incubate |
Revision as of 17:54, 13 October 2013
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10/10/12Site Directed Mutagenesis-Removing the BSMBI cut site from BD002 reporter gene
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