IGEM:Harvard/2008/Lab Notebooks/DailyBook/Week13/Chemical and Light

Colony PCR 09/30

Should grow only on carb

• 1-30: 20, 23, 25 (picked all three) --> P38+130
• 31-60: 31-60 (picked 31, 35, 39, 42, 46, 50, 55, 60) --> P39+130

Should grow only on kan

• 61-90: none --> P116+130
• 91-120: none --> P117+130
• 121-150: none --> P108+130

--Upload picture of colony PCR gel--

Picked colony 20 (P38+130) and 31, 39, 60 (P39+130) and grew them up in liquid cultures (LB Amp).

Amy's mutants

RE digests 10/01

Numbers refer to the master plate from 09/30 colony PCR. We digested with EcoRI, SpeI, and XmnI.

• P38+130 #20
• P39+130 #31
• P39+130 #39

The expected band sizes are 2.2 kb (part), 1.6 kb and 0.4 kb (both vector backbone).

Ligations/transformations 10/03

We ligated the following plasmids. The ligations were transformed into XL1-Blue.

• P108>131
• P38+51>131
• P122>131
• P38+130>101
• P130>133
• P130>134

There were two colonies on the P130>134 plate. These were grown up in liquid culture to be sent for sequencing. There were no colonies on the other plates.

Shewanella transformations 10/05

We ethanol precipitated the above ligations prior to electroporation of Shewanella.

We transformed the following plasmids into wildtype Shewanella:

• P130>134
• P38+51>131
• P130>133
• P122>131
• P38+130>101
• P108>131
• P138 #1
• P139 #1
• P139 #2
• P139 #3
• P139 #4
• P139 #5
• P140
• P142

We transformed the following plasmids into ΔmtrB Shewanella:

• P130>134
• P38+51>131
• P130>133
• P122>131
• P38+130>101
• P108>131
• P138 #2
• P139 #1
• P140
• P142