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 chromasome 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^