IGEM:Harvard/2006/Cyanobacteria/Notebook/2006-9-29

Digestions

We need to digest:

• J36012 (KaiA)\SP + J36010 ( KaiC)\XP
• J36011 (KaiB)\SP + J36010 (KaiC)\XP
• B0034 \XP + J36000 KaiC \SP
• B0034 \XP + J36001 KaiB \SP

(Note: Just KaiA, B, or C on a backbone we can submit the geneart plasmids; as far as i can tell they do not have other E/X/S/P sites)

Then:

J36010 \XP: 8 lanes J36011 \SP: 4 lanes J36012 \SP: 4 lanes B0034 \XP: 8 lanes J36000 \SP: 4 lanes J36001 \SP: 4 lanes

Protocols:

B0034 \XP: 8 samples

1. • 7.6 uL DNA (62.32)
   • 1 µL 10 x BSA (8.2)
   • 1 µL 10x buffer #3 (8.2)
   • 0.2 µL Xba1 (1.64)
   • 0.2 µL Pst1 (1.64)

J36010 \XP: (8 samples)

1. • 7.6 uL DNA (62.32)
   • 1 µL 10 x BSA (8.2)
   • 1 µL 10x buffer #3 (8.2)
   • 0.2 µL Xba1 (1.64)
   • 0.2 µL Pst1 (1.64)

J36011 \SP: (4 samples)

1. • 5 uL DNA (20.5)
   • 1 µL 10 x BSA (4.1)
   • 1 µL 10x buffer #2 (4.1)
   • 0.2 µL Spe1 (.82)
   • 0.2 µL Pst1 (.82)
   • 2.6 uL H20 (10.66)

J36012 \SP: (4 samples)

1. • 7.6 uL DNA (31.16)
   • 1 µL 10 x BSA (4.1)
   • 1 µL 10x buffer #2 (4.1)
   • 0.2 µL Spe1 (.82)
   • 0.2 µL Pst1 (.82)

J36001 \SP: (4 samples)

1. • 7.6 uL DNA (31.16)
   • 1 µL 10 x BSA (4.1)
   • 1 µL 10x buffer #2 (4.1)
   • 0.2 µL Spe1 (.82)
   • 0.2 µL Pst1 (.82)

J36000 (KaiC \SP):

1. • 4 uL DNA (16.4)
   • 1 µL 10 x BSA (4.1)
   • 1 µL 10x buffer #2 (4.1)
   • 0.2 µL Spe1 (.82)
   • 0.2 µL Pst1 (.82)
   • 3.6 uL H20 (14.76)

16h @ 45C