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"|<html><img src="/images/9/94/Report.png" border="0" /></html> [[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>[[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]]<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>}}
|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]]&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}}
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| bgcolor=#cfcfcf | Reagent
| bgcolor=#cfcfcf | Reagent
| bgcolor=#cfcfcf | Volume
| bgcolor=#cfcfcf | Volume
| rowspan="7" | Expected:<br>1,2. DPRE (E/S) = ###, 3200<br>3. BD004 (E/X) = 7318
| rowspan="7" | Expected:<br>1,2. DPRE (E/S) = 1740*, 3200<br>3. BD004 (E/X) = 7316*<br>*Bands cut & purified
| rowspan="7" | [[Image:KAH032015_gel1.jpg|270px|Hover name]]<br>30 μL/lane, 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
| 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]
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| DNA (plasmid) || 20.0 μL
| DNA (plasmid) || 20.0 μL
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| enzyme 2 || 1.0
| enzyme 2 || 1.0
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| dH<sub>2</sub>O || ---
| dH<sub>2</sub>O || 5.0
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| &nbsp; || 30 μL --> 37°C/ ~30 min.
| &nbsp; || 30 μL --> 37°C/ ~30 min.
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* Conclusions
* Conclusions
** tba
** All look good
 


* Miniprep conc.'s
* Miniprep conc.'s
** Combined the pairs of preps. Pairs look identical.
{| {{table}} cellspacing="3" <!-- [DNA] table -->
{| {{table}} cellspacing="3" <!-- [DNA] table -->
|- bgcolor=#cfcfcf  
|- bgcolor=#cfcfcf  
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* 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

Project name Main project page
Previous entry      Next entry

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

  • ---Karmella 15:12, 27 March 2015 (EDT):
  • Sigma GenElute Miniprep kit.
  • Elute with 70μL elution buffer
  1. DPRE a
  2. DPRE b
  • Digests (Fermentas FD)
    • E/S to prep for insertion upstream of BD004
    • Also digest BD004 w/ E/X
Reagent Volume Expected:
1,2. DPRE (E/S) = 1740*, 3200
3. BD004 (E/X) = 7316*
*Bands cut & purified 		Hover name
30 μL/lane, 1% agarose; Ladder
DNA (plasmid) 20.0 μL
10x buffer 3.0
enzyme 1 1.0
enzyme 2 1.0
dH2O 5.0
  30 μL --> 37°C/ ~30 min.


  • Conclusions
    • All look good


  • Miniprep conc.'s
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
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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