Team:Paris/August 9

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Contents

1 Ligation Day

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

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