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

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

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

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^