Team:Paris/August 9

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{{Paris/Calendar_Links|August 8|August 10}}
{{Paris/Calendar_Links|August 8|August 10}}
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=='''Ligation Day'''==
 +
==='''List of ligations'''===
 +
{| Border="2"
 +
|align="center"|'''Ligation name'''
 +
|align="center"|'''Insert name '''
 +
|align="center"|'''Vector name'''
 +
|-
 +
|align="center"|L 132
 +
|align="center"|D 139 (flhDC (gene))
 +
|align="center"|D 142 (pSB1A2)
 +
|-
 +
|align="center"|L 133
 +
|align="center"|D 140 (OmpR*)
 +
|align="center"|D 142 (pSB1A2)
 +
|-
 +
|align="center"|L 134
 +
|align="center"|D 141 (EnvZ*)
 +
|align="center"|D 142 (pSB1A2)
 +
|-
 +
|align="center"|L 135
 +
|align="center"|D 133 (pflgA)
 +
|align="center"|D 137 (pSB3K3)
 +
|-
 +
|align="center"|L 136
 +
|align="center"|D 134 (pflgB)
 +
|align="center"|D 137 (pSB3K3)
 +
|-
 +
|align="center"|L 137
 +
|align="center"|D 135 (pflhB)
 +
|align="center"|D 137 (pSB3K3)
 +
|-
 +
|align="center"|L 138
 +
|align="center"|D 138 (E0240)
 +
|align="center"|D 137 (pSB3K3)
 +
|}
 +
 +
 +
 +
==='''Measure of DNA concentration'''===
 +
First of all, we have to determine the concentration of DNA of the different templates.
 +
 +
We used the biophotometer.
 +
*Sample: 5µL of template DNA + 95µL of water
 +
*Blank: 5µL of EB buffer + 95 µL of water
 +
 +
{| Border="2"
 +
|align="center"|'''Ligation name'''
 +
|align="center"|'''[DNA] diluted<br>ng/µL'''
 +
|align="center"|'''[DNA] template<br>ng/µL'''
 +
|-
 +
|align="center"|D 132
 +
|align="center"| 1
 +
|align="center"| 20
 +
|-
 +
|align="center"|D 133
 +
|align="center"| 1
 +
|align="center"| 20
 +
|-
 +
|align="center"|D 134
 +
|align="center"| 1
 +
|align="center"| 20
 +
|-
 +
|align="center"|D 137
 +
|align="center"| 1
 +
|align="center"| 20
 +
|-
 +
|align="center"|D 138
 +
|align="center"| 1
 +
|align="center"| 20
 +
|-
 +
|align="center"|D 139
 +
|align="center"| 0
 +
|align="center"| 0
 +
|-
 +
|align="center"|D 140
 +
|align="center"| 1
 +
|align="center"| 20
 +
|-
 +
|align="center"|D 141
 +
|align="center"| 2
 +
|align="center"| 40
 +
|-
 +
|align="center"|D 142
 +
|align="center"| 1
 +
|align="center"| 20
 +
|}
 +
==='''Protocol of the ligation'''===
 +
* 2 µL T4 Ligase Buffer 10X
 +
* X µg/µL vector
 +
* 3 or 4 x X µg/µL insert
 +
* Pure water qsp 20 µL
 +
* 1 µL T4 ligase
 +
* O/N at 16°C + sunday 16°C
 +
 +
{| Border="2"
 +
|align="center"|'''Ligation name'''
 +
|align="center"|'''Insert Volume'''
 +
|align="center"|'''Vector Volume'''
 +
|-
 +
|align="center"|L 132
 +
|align="center"|3 µL
 +
|align="center"|5 µL
 +
|-
 +
|align="center"|L 133
 +
|align="center"|3 µL
 +
|align="center"|5 µL
 +
|-
 +
|align="center"|L 134
 +
|align="center"|3 µL
 +
|align="center"|5 µL
 +
|-
 +
|align="center"|L 135
 +
|align="center"|2.5 µL
 +
|align="center"|2.5 µL
 +
|-
 +
|align="center"|L 136
 +
|align="center"|2.5 µL
 +
|align="center"|2.5 µL
 +
|-
 +
|align="center"|L 137
 +
|align="center"|2.5 µL
 +
|align="center"|2.5 µL
 +
|-
 +
|align="center"|L 138
 +
|align="center"|2.5 µL
 +
|align="center"|2.5 µL
 +
|}

Latest revision as of 17:11, 13 August 2008

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Contents

Ligation Day

List of ligations

Ligation name Insert name Vector name
L 132 D 139 (flhDC (gene)) D 142 (pSB1A2)
L 133 D 140 (OmpR*) D 142 (pSB1A2)
L 134 D 141 (EnvZ*) D 142 (pSB1A2)
L 135 D 133 (pflgA) D 137 (pSB3K3)
L 136 D 134 (pflgB) D 137 (pSB3K3)
L 137 D 135 (pflhB) D 137 (pSB3K3)
L 138 D 138 (E0240) D 137 (pSB3K3)


Measure of DNA concentration

First of all, we have to determine the concentration of DNA of the different templates.

We used the biophotometer.

  • Sample: 5µL of template DNA + 95µL of water
  • Blank: 5µL of EB buffer + 95 µL of water
Ligation name [DNA] diluted
ng/µL 		[DNA] template
ng/µL
D 132 1 20
D 133 1 20
D 134 1 20
D 137 1 20
D 138 1 20
D 139 0 0
D 140 1 20
D 141 2 40
D 142 1 20

Protocol of the ligation

  • 2 µL T4 Ligase Buffer 10X
  • X µg/µL vector
  • 3 or 4 x X µg/µL insert
  • Pure water qsp 20 µL
  • 1 µL T4 ligase
  • O/N at 16°C + sunday 16°C
Ligation name Insert Volume Vector Volume
L 132 3 µL 5 µL
L 133 3 µL 5 µL
L 134 3 µL 5 µL
L 135 2.5 µL 2.5 µL
L 136 2.5 µL 2.5 µL
L 137 2.5 µL 2.5 µL
L 138 2.5 µL 2.5 µL
Retrieved from "http://2008.igem.org/Team:Paris/August_9"