User:Karmella Haynes/Notebook/BioBrick cloning/2013/03/06

03/06/13

• Colony PCR: for Rene's project

Colony PCR

• Streaked single colonies on gridded plates yesterday
• Used bit of streak for PCR today
• All parts are on pSB1A3-based vectors (http://partsregistry.org/Part:pSB1A3)
  • Use primers VF2 and VR to amplify inserts

Reagent	Volume	Expected: Primers add ~250bp 1-3. K084007 (835 bp) = 1085 4-6. R0071 (53 bp) = 303 7. K.. (# bp) = ##	10 μL/lane; 1% agarose; Ladder
Bacteria from tip	---
10 μM VF2 primer	1.0
10 μM VR primer	1.0
2x GoTaq Green	12.5
dH2O	10.5
	25.0

PCR

• 95°C, 3 min
• [95°C, 15 sec; 57°C 15 sec; 72 °C 60 sec] x35
• 72°C, 3 min
• 4°C, ∞