Haynes Lab:Notebook/Jan/2015/03/28: Difference between revisions

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** Elute & back-elute w/ 70 μL elution sln.
** Elute & back-elute w/ 70 μL elution sln.
# BD006/MV2 plate 1, a
# BD006/MV2 plate 1, a
# BD006/MV2 plate 1, b
# BD006/MV2 plate 2, a
# BD006/MV2 plate 2, b




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| bgcolor=#cfcfcf | Reagent
| bgcolor=#cfcfcf | Reagent
| bgcolor=#cfcfcf | Volume
| bgcolor=#cfcfcf | Volume
| rowspan="7" | Expected:<br>* BD006/MV2 = 8482, 3030, 32<br>* MV2 only = 4273, 3030, 32
| rowspan="7" | Expected:<br>* BD006/MV2 = 7207, 1243, 288<br>* MV2 only = 2998, 1243, 288<br>* BD006/V0120 = 7036, 497
| 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]
| rowspan="7" | [[Image:KAH032815_gel1.jpg|270px|Hover name]]<br>30 μL/lane, 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
|-
|-
| DNA (plasmid) || 5.0 μL
| DNA (plasmid) || 3.0 μL
|-
|-
| 10x buffer || 1.5
| 10x buffer || 1.5
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| ApaLI || 1.0
| ApaLI || 1.0
|-
|-
| dH<sub>2</sub>O || 7.5
| dH<sub>2</sub>O || 9.5
|-
|-
| &nbsp; || 15 μL --> 37°C/ ~10 min.
| &nbsp; || 15 μL --> 37°C/ ~10 min.
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* Conclusions
* Conclusions
** tba
** Success! All four look like BD006/MV2
** Submit for sequencing next week
 
 
* Miniprep conc.'s
{| {{table}} cellspacing="3" <!-- [DNA] table -->
|- bgcolor=#cfcfcf
| Sample || OD260 || 260/280 || ng/μL
|-
| 1. BD006/MV2 1-a || 0.274 || 1.44 || 274.10
|-
| 2. BD006/MV2 1-b || 0.299 || 1.93 || 298.68
|-
| 3. BD006/MV2 2-a || 0.331 || 1.93 || 330.69
|-
| 4. BD006/MV2 2-b || 0.306 || 1.90 || 305.94
|}




Revision as of 13:09, 28 March 2015

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03/28/15

  • Minipreps - BD006/MV2 colonies (from 3/26/15 ligations), 5 mL each (4 total)


Minipreps - BD006/MV2 colonies

  • Sigma GenElute kit.
    • Elute & back-elute w/ 70 μL elution sln.
  1. BD006/MV2 plate 1, a
  2. BD006/MV2 plate 1, b
  3. BD006/MV2 plate 2, a
  4. BD006/MV2 plate 2, b


  • Restriction Digest check (Fermentas FD)
    • Cut with ApaLI (E/S insertion got rid of BB RFC23 cut sites)


Reagent Volume Expected:
* BD006/MV2 = 7207, 1243, 288
* MV2 only = 2998, 1243, 288
* BD006/V0120 = 7036, 497 		Hover name
30 μL/lane, 1% agarose; Ladder
DNA (plasmid) 3.0 μL
10x buffer 1.5
ApaLI 1.0
dH2O 9.5
  15 μL --> 37°C/ ~10 min.


  • Conclusions
    • Success! All four look like BD006/MV2
    • Submit for sequencing next week


  • Miniprep conc.'s
Sample OD260 260/280 ng/μL
1. BD006/MV2 1-a 0.274 1.44 274.10
2. BD006/MV2 1-b 0.299 1.93 298.68
3. BD006/MV2 2-a 0.331 1.93 330.69
4. BD006/MV2 2-b 0.306 1.90 305.94



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