IGEM:Peking/2007/Count-Conjugation-Notebook/2007-7-18

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Contents

Miniprep

  • Miniprep & preservation: I7100.

Digestion Test: I7100 and E0040

  • I7100 and E0040.
  • EcoR I single digestion.
  • Digestion system contains (totally 20uL): 2uL 10*H, 0.5uL EcoR I, 3uL plasmid, 15uL ddH2O.

electrophoresis result

  • from Left to Right:
  1. I7100 @ EcoRI
  2. E0040 @ EcoRI
  3. I7100 plasmid
  4. E0040 plasmid
  5. Marker

Image:2007-7-18_I7100_E0040_EcoRI.jpg‎

Double Digesting E0040 and I7100 for ligation

  • Method choice test :Oven or Water bath
  • Digesting E0040 and I7100 with EcoR1/SpeI and EcoRI/XbaI.
  • Digestion system contains (totally 20uL):
  1. 3uL 10*H, 1uL EcoRI, 1uL SpeI, 6uL plasmid, 10uL ddH2O.
  2. 3uL 10*M, 1uL EcoRI, 1uL XbaI, 6uL plasmid, 10uL ddH2O.
  • 37℃ culutre for 12 hours.

electrophoresis result

  • from Left to Right:
  1. oven:I7100 @ EcoRI/XbaI
  2. Oven:E0040 @ EcoRI/SpeI
  3. Water Bath: I7100 @ EcoRI/XbaI
  4. Water Bath: E0040 @ EcoRI/SpeI
  5. Marker

Image:I7100-EcoRI+XbaI_E0040-EcoRI+SpeI.jpg

conclusion

Water bath is better.

OriT Knock Out

  • By Xu Anting and Liu Ting

Chromosome Extraction

  • Received chromosome of F/R751/pSC101 strain and diluted in 1:50 ddH2O (pH 8.0).

Travel to Grassland!

  • Everyone at Room 307 in Biotech Building planned to have a travel for three days. All experiment suspended ^o^
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