Haynes Lab:Notebook/Jan/2015/03/27: Difference between revisions
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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span> | |style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span> | ||
|style="background-color: #F2F2F2" align="center"| | |style="background-color: #F2F2F2" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]] }}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}} | ||
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== | ==03/27/15== | ||
* | * Cultures - BD006/MV2 colonies (from 3/26/15 ligations), 5 mL each (4 total) | ||
* Plasmids - DPRE minipreps (5 mL cultures) | |||
---- | |||
'''Plasmids - DPRE minipreps''' | |||
*'''[[User:Karmella Haynes|---Karmella]] 15:12, 27 March 2015 (EDT)''': | |||
* Sigma GenElute Miniprep kit. | |||
* Elute with 70μL elution buffer | |||
# DPRE a | |||
# DPRE b | |||
* Digests (Fermentas FD) | |||
** E/S to prep for insertion upstream of BD004 | |||
** Also digest BD004 w/ E/X | |||
{| {{table}} cellspacing="3" <!-- Digest rxn. table --> | |||
|- valign="top" | |||
| bgcolor=#cfcfcf | Reagent | |||
| bgcolor=#cfcfcf | Volume | |||
| rowspan="7" | Expected:<br>1,2. DPRE (E/S) = 1740*, 3200<br>3. BD004 (E/X) = 7316*<br>*Bands cut & purified | |||
| rowspan="7" | [[Image:KAH032715_gel1.jpg|270px|Hover name]]<br>30 μL/lane, 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder] | |||
|- | |||
| DNA (plasmid) || 20.0 μL | |||
|- | |||
| 10x buffer || 3.0 | |||
|- | |||
| enzyme 1 || 1.0 | |||
|- | |||
| enzyme 2 || 1.0 | |||
|- | |||
| dH<sub>2</sub>O || 5.0 | |||
|- | |||
| || 30 μL --> 37°C/ ~30 min. | |||
|} | |||
* Conclusions | |||
** All look good | |||
* Miniprep conc.'s | |||
{| {{table}} cellspacing="3" <!-- [DNA] table --> | |||
|- bgcolor=#cfcfcf | |||
| Sample || OD260 || 260/280 || ng/μL | |||
|- | |||
| 1. DPRE a mp || 0.411 || 1.89 || 411.47 | |||
|- | |||
| 2. DPRE b mp || 0.411 || 1.89 || 410.51 | |||
|} | |||
* Gel purification conc.'s | |||
** Sigma gel extraction kit, eluted & back-eluted w/ 30 μL elution sln | |||
{| {{table}} cellspacing="3" <!-- [DNA] table --> | |||
|- bgcolor=#cfcfcf | |||
| Sample || OD260 || 260/280 || ng/μL | |||
|- | |||
| 1. DPRE a (E/S) || 0.017 || 1.88 || 16.81 | |||
|- | |||
| 2. DPRE b (E/S) || 0.013 || 1.54 || 13.41 | |||
|- | |||
| 2. BD004 (E/X) || 0.006 || 1.85 || 6.46 | |||
|} | |||
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Latest revision as of 00:52, 27 September 2017
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03/27/15
Plasmids - DPRE minipreps
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