Team:Paris/August 18

From 2008.igem.org

(Difference between revisions)
(→Analysis of the cloning of OmpR and EnvZ)
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{{Paris/Calendar_Links|August 17|August 19}}
{{Paris/Calendar_Links|August 17|August 19}}
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=Analysis of the cloning of OmpR* and EnvZ*=
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=Analysis of the cloning of OmpR and EnvZ=
==PCR screening==
==PCR screening==
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*0,5 µL O19
*0,5 µL O19
*12 µL water
*12 µL water
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We wanted to analyse the transformants of the ligations L133 (OmpR* in pSB1A2) and L134 (EnvZ* in pSB1A2) from the experience of august 9th/11th.
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We wanted to analyse the transformants of the ligations L133 (OmpR in pSB1A2) and L134 (EnvZ in pSB1A2) from the experience of august 9th/11th.
<br>3 clones of L133 and 3 clones of L134 were analysed.
<br>3 clones of L133 and 3 clones of L134 were analysed.
<br>Each clone tested was then cultured in 7,5 mL LB + ampicilline.
<br>Each clone tested was then cultured in 7,5 mL LB + ampicilline.
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|align="center"|XbaI - PstI
|align="center"|XbaI - PstI
|align="center"|5.81 µL
|align="center"|5.81 µL
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|}
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 +
=='''Ligation'''==
 +
{|border="1" style="text-align: center"
 +
|'''Ligation Name'''
 +
|'''Vector Name'''
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|'''Volume Vector (µL)'''
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|'''Insert'''
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|'''Volume Insert (µL)'''
 +
|-
 +
|L153
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|D123.1
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|1.54
 +
|D165
 +
|1.69
 +
|-
 +
|L154
 +
|D103.2
 +
|1.14
 +
|D165
 +
|1.68
|}
|}

Revision as of 15:38, 19 August 2008

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Contents

Analysis of the cloning of OmpR and EnvZ

PCR screening

  • positive control: S158 (pSB3K3)
  • negative control: no template
  • screening PCR programm
  • elongation time: 1 min 30
  • primers: O18 & O19

Reaction mixture (25 µL total volume)

  • 12,5 µL Quick load PCR Mix
  • 0,5 µL O18
  • 0,5 µL O19
  • 12 µL water

We wanted to analyse the transformants of the ligations L133 (OmpR in pSB1A2) and L134 (EnvZ in pSB1A2) from the experience of august 9th/11th.
3 clones of L133 and 3 clones of L134 were analysed.
Each clone tested was then cultured in 7,5 mL LB + ampicilline.


Miniprep and stock glycerol of L151

Stock number Ligation name Biobricks Description
S162.1 L151.1 Part icon rbs.pngIcon coding.pngPart icon terminator.pngPart icon terminator.png rbs-LasR - Double terminator
S162.2 L151.2
S162.3 L151.3


Miniprep Name Ligation name Antibio Biobricks Description
MP162.1 L151.1 Kan Part icon rbs.pngIcon coding.pngPart icon terminator.pngPart icon terminator.png rbs-LasR - Double terminator
MP162.2 L151.2
MP162.3 L151.3

Digestion of MP162

Measurement of the concentration of MPs 162

Miniprep Name Concentration (µg/mL) Ratio 260/280
MP162.1 151 1.71
MP162.2 172 1.73
MP162.3 149 1.73

Digestion

protocol

Digestion name Template DNA Enzymes Volume of DNA
D165 MP162.2 XbaI - PstI 5.81 µL

Ligation

Ligation Name Vector Name Volume Vector (µL) Insert Volume Insert (µL)
L153 D123.1 1.54 D165 1.69
L154 D103.2 1.14 D165 1.68
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