User:Wesley J. Houston/Notebook/MAP/2013/03/08

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(3/8/13)
Current revision (01:10, 26 March 2013) (view source)
(3/8/13)
 
Line 13: Line 13:
> Ligations
> Ligations
 +
{| class="wikitable" border="0" cellspacing="3" <!-- Ligation rxn table -->
 +
| &nbsp;            || 1    || 2    ||
 +
|-
 +
| Insert DNA (KAH182)        || 3.0  || ---  ||
 +
|-
 +
| Vector DNA (KAH201)        || 0.5  || 0.5  ||
 +
|-
 +
| 2x lgn buf (Roche) || 5.0  || 5.0  ||
 +
|-
 +
| T4 ligase (NEB)    || 1.0  || 1.0  ||
 +
|-
 +
| dH<sub>2</sub>O    ||  0.5 ||  3.5 ||
 +
|-
 +
| &nbsp;            || 10 μL || 10 μL ||
 +
|}
 +
 +
* Follow quick transformation protocol
 +
* Add 30 uL DH5α-Turbo to each ligation; plate on 100 μg/mL Amp agar
 +
{| class="wikitable" border="0" cellspacing="3" <!-- Ligations table -->
{| class="wikitable" border="0" cellspacing="3" <!-- Ligations table -->
| <u>Ligation</u> || <font color="blue"><u>Plate results (lig : neg crtl)</u> 03/08/13</font>
| <u>Ligation</u> || <font color="blue"><u>Plate results (lig : neg crtl)</u> 03/08/13</font>

Current revision

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3/8/13

  • Ligation was repeated.
  • Attempted to grow more liquid cultures of ligated plasmids.
  • Attempted to confirm ligation by streaking the colonies on an agar plate.
  • All attempts proved unsuccessful.

> Ligations

  1 2
Insert DNA (KAH182) 3.0 ---
Vector DNA (KAH201) 0.5 0.5
2x lgn buf (Roche) 5.0 5.0
T4 ligase (NEB) 1.0 1.0
dH2O 0.5 3.5
  10 μL 10 μL
  • Follow quick transformation protocol
  • Add 30 uL DH5α-Turbo to each ligation; plate on 100 μg/mL Amp agar
Ligation Plate results (lig : neg crtl) 03/08/13
1. KAH201 (S/P)/3403, 25 ng + KAH182 (X/P)/1675, 26 ng 0
2. KAH201 (S/P)/3403, 25 ng 0
  • No colonies were present in the ligation plate.


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