Jessica Karen Wong/Notebook/2007-7-7

• Took out overnight transformation plates
  • both the sequential and double digests of T9002 and 3K3 had lots of colonies
  • E0240-3K3 had lots of colonies, E0240-1AK3 had two
  • I2055-1AK3 had lots of colonies, I-1AT3 and I-1AC3 had none

• Made cell suspensions of 4 T-3K3 colonies, 4 I-1AK3 colonies, 4 E-3K3, and 2 E-1AK3
• Did a 10 ul colony PCR on each cell suspensions
  • Used 2min elongation time because T9002 is 2261bp b/t VF and VR

• Ran gel of I2055 and got a bright band just under 1kb
  • Not sure if it contains promoter or not
• PCR cleaned I2055
• Double digested I2055 with Mfe1 and Nsi1
  • Used buffer 2 and 4ul DNA

• Overnight liquid cultures of I2056 still didn't grow (maybe lost Amp resistance?)
• Made new overnights of I2056 containing only Tet

• Did a 2-way ligation of T9002 and 1AK3
• Transformed 3ul of the ligation and plated on Kan