Magni:Back Door

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== Reagents ==
+
== Antibiotics ==
 +
*Ampicillin - 100 mg/ml in deionized water (-20°C)
 +
*Chloramphenicol - 34 mg/ml in ethanol (-20°C)
 +
*Kanamycin - 50 mg/ml in deionized water (-20°C)
 +
 
 +
== Inducers and substrates ==
 +
*HSL (3-oxo-C6-homoserine lactone) - 2 mM in water (-20°C)
 +
*aTc (Anhydrotetracycline) - 2 mg/ml in ethanol 70% or 100 ug/ml in ethanol 70% (-20°C) LIGHT SENSITIVE
 +
*IPTG Ready Made - 200 mM (-20°C)
 +
*X-Gal - 40 mg/ml in DMF (-20°C)
== ''E. coli'' strains ==
== ''E. coli'' strains ==
 +
*BW20767 (Registry)
 +
 +
*BW23474 (CGSC)
 +
**pir-116 strain, used to propagate the R6K conditional replication origin at high copy
 +
 +
*BW25141 (CGSC)
 +
**pir+ strain, used to propagate the R6K conditional replication origin at medium copy
 +
 +
*BW25142 (CGSC)
 +
**pir-116 strain, used to propagate the R6K conditional replication origin at high copy (equivalent to BW23474)
 +
 +
*CW2553 + pJat8 (Registry)
 +
 +
*DB3.1 (Invitrogen)
 +
**can propagate plasmids with ccdB toxin
 +
 +
*DH5alpha (Invitrogen)
 +
**general purpose cloning
 +
**blue/white ok
 +
 +
*KRX (Promega)
 +
**overexpresses LacI via the lacIq mutation on the F plasmid
 +
**can be used for autoinduction protocol (see Promega handbook)
 +
**blue/white ok
 +
 +
*MC1061 (CGSC)
 +
**no blue/white screening
 +
 +
*MG1655 (CGSC)
 +
**K-12 sequenced strain
 +
**no blue/white screening
 +
 +
*MG1655-Z1 (Univ of Washington)
 +
**MG1655 with lacI- and tetR- overexpression cassettes in the genome, in the attB(lambda) locus
 +
 +
*TOP10 (Invitrogen)
 +
**general purpose cloning
 +
**blue/white ok
 +
 +
*XL1-Blue (Stratagene)
== Important glycerol stocks ==
== Important glycerol stocks ==
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Revision as of 09:09, 30 April 2012

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Contents

Antibiotics

  • Ampicillin - 100 mg/ml in deionized water (-20°C)
  • Chloramphenicol - 34 mg/ml in ethanol (-20°C)
  • Kanamycin - 50 mg/ml in deionized water (-20°C)

Inducers and substrates

  • HSL (3-oxo-C6-homoserine lactone) - 2 mM in water (-20°C)
  • aTc (Anhydrotetracycline) - 2 mg/ml in ethanol 70% or 100 ug/ml in ethanol 70% (-20°C) LIGHT SENSITIVE
  • IPTG Ready Made - 200 mM (-20°C)
  • X-Gal - 40 mg/ml in DMF (-20°C)

E. coli strains

  • BW20767 (Registry)
  • BW23474 (CGSC)
    • pir-116 strain, used to propagate the R6K conditional replication origin at high copy
  • BW25141 (CGSC)
    • pir+ strain, used to propagate the R6K conditional replication origin at medium copy
  • BW25142 (CGSC)
    • pir-116 strain, used to propagate the R6K conditional replication origin at high copy (equivalent to BW23474)
  • CW2553 + pJat8 (Registry)
  • DB3.1 (Invitrogen)
    • can propagate plasmids with ccdB toxin
  • DH5alpha (Invitrogen)
    • general purpose cloning
    • blue/white ok
  • KRX (Promega)
    • overexpresses LacI via the lacIq mutation on the F plasmid
    • can be used for autoinduction protocol (see Promega handbook)
    • blue/white ok
  • MC1061 (CGSC)
    • no blue/white screening
  • MG1655 (CGSC)
    • K-12 sequenced strain
    • no blue/white screening
  • MG1655-Z1 (Univ of Washington)
    • MG1655 with lacI- and tetR- overexpression cassettes in the genome, in the attB(lambda) locus
  • TOP10 (Invitrogen)
    • general purpose cloning
    • blue/white ok
  • XL1-Blue (Stratagene)

Important glycerol stocks

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