User:Wesley J. Houston/Notebook/MAP/2013/04/01: Difference between revisions

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(Autocreate 2013/04/01 Entry for User:Wesley_J._Houston/Notebook/MAP)
 
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==Entry title==
==4/1/13==
* Insert content here...
* Completed 2 ligations of KAH201 (S/P)/3403 + KAH182 (X/P)/1675
** First ligation used KAH201 (S/P)/3403 and KAH182 (X/P) 1675 made on [http://openwetware.org/wiki/User:Wesley_J._Houston/Notebook/MAP/2013/03/26 3/26/13]
** Second ligation used the same KAH182 (X/P)/1675 and KAH201 (S/P)/3403 made on [http://openwetware.org/wiki/User:Wesley_J._Houston/Notebook/MAP/2013/02/08 2/8/13]


> Ligation 1
{| class="wikitable" border="0" cellspacing="3" <!-- Ligation rxn table -->
| &nbsp;            || 1    || 2    ||
|-
| Insert DNA (KAH182)        || 3.5  || ---  ||
|-
| Vector DNA (KAH201)        || 1.0  || 1.0  ||
|-
| 2x lgn buf (Roche) || 5.5  || 5.5  ||
|-
| T4 ligase (NEB)    || 1.0  || 1.0  ||
|-
| dH<sub>2</sub>O    ||  0.0 ||  3.5 ||
|-
| &nbsp;            || 11 μL || 11 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 -->
| <u>Ligation</u> || <font color="blue"><u>Plate results (lig : neg crtl)</u>04/01/13: Ligation 1</font>
|-
| 1. KAH201 (S/P)/<font color="blue">3403</font>, 25 ng + KAH182 (X/P)/1675, 26 ng || <font color="blue">0</font>
|-
| 2. KAH201 (S/P)/3403, 25 ng || 0
|}
> Ligation 2
{| class="wikitable" border="0" cellspacing="3" <!-- Ligation rxn table -->
| &nbsp;            || 1    || 2    ||
|-
| Insert DNA (KAH182)        || 3.5  || ---  ||
|-
| Vector DNA (KAH201)        || 0.7  || 0.7  ||
|-
| 2x lgn buf (Roche) || 5.2  || 5.2  ||
|-
| T4 ligase (NEB)    || 1.0  || 1.0  ||
|-
| dH<sub>2</sub>O    ||  0.6 ||  4.1 ||
|-
| &nbsp;            || 11 μL || 11 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 -->
| <u>Ligation</u> || <font color="blue"><u>Plate results (lig : neg crtl)</u>04/01/13: Ligation 2</font>
|-
| 1. KAH201 (S/P)/<font color="blue">3403</font>, 25 ng + KAH182 (X/P)/1675, 26 ng || <font color="blue">~40</font>
|-
| 2. KAH201 (S/P)/3403, 25 ng || 0
|}
* Ligation 2 appears successful, with ~40 colonies in the ligation plate and 0 colonies in the negative control plate. [[Image:4.1.13ligation.jpg‎ |150px|Results of the ligation]]
** The next step will be to confirm the assembly, to be completed on [http://openwetware.org/wiki/User:Wesley_J._Houston/Notebook/MAP/2013/04/04 4/4/13]


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Revision as of 18:20, 2 April 2013

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4/1/13

  • Completed 2 ligations of KAH201 (S/P)/3403 + KAH182 (X/P)/1675
    • First ligation used KAH201 (S/P)/3403 and KAH182 (X/P) 1675 made on 3/26/13
    • Second ligation used the same KAH182 (X/P)/1675 and KAH201 (S/P)/3403 made on 2/8/13

> Ligation 1

  1 2
Insert DNA (KAH182) 3.5 ---
Vector DNA (KAH201) 1.0 1.0
2x lgn buf (Roche) 5.5 5.5
T4 ligase (NEB) 1.0 1.0
dH2O 0.0 3.5
  11 μL 11 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)04/01/13: Ligation 1
1. KAH201 (S/P)/3403, 25 ng + KAH182 (X/P)/1675, 26 ng 0
2. KAH201 (S/P)/3403, 25 ng 0


> Ligation 2

  1 2
Insert DNA (KAH182) 3.5 ---
Vector DNA (KAH201) 0.7 0.7
2x lgn buf (Roche) 5.2 5.2
T4 ligase (NEB) 1.0 1.0
dH2O 0.6 4.1
  11 μL 11 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)04/01/13: Ligation 2
1. KAH201 (S/P)/3403, 25 ng + KAH182 (X/P)/1675, 26 ng ~40
2. KAH201 (S/P)/3403, 25 ng 0
  • Ligation 2 appears successful, with ~40 colonies in the ligation plate and 0 colonies in the negative control plate. Results of the ligation
    • The next step will be to confirm the assembly, to be completed on 4/4/13


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