IGEM:MIT/2007/Notebook/2007-6-14

From OpenWetWare

(Difference between revisions)
Jump to: navigation, search
Line 13: Line 13:
**Analyze plates
**Analyze plates
**innoculate 4ml LB/Amp
**innoculate 4ml LB/Amp
 +
 +
==Procedures==
 +
 +
Regular cell membranes are not permeable to plasmids, so we much use ions (CaCl) to make the bacteria competent and able to accept foreign DNA.
 +
Start: 2 vials of competent bacteria (500 uL each)
 +
9 Transformations
 +
U V (2 uL)
 +
dV (5 uL)
 +
dV + dI (5 uL)
 +
ddV (5 uL)
 +
ddV + ddI (5 uL)
 +
dV (10 uL)
 +
dV + dI (10 uL)
 +
ddV (10 uL)
 +
ddV + ddI (10 uL)
 +
 +
#Aliquot 5 uL and 10 uL of ligation reactions into eppendorfs
 +
#Pipette 50 uL of competent DH5a

Revision as of 09:40, 14 June 2007

Grad Advisors

  • Morning: Debbie
  • Afternoon: Brian

LAB WORK

  • Make competent E. coli cells
  • Transformation into competent cells
  • Plate transformed cells


Depending on how fast cells grow......

  • Night time
    • Analyze plates
    • innoculate 4ml LB/Amp

Procedures

Regular cell membranes are not permeable to plasmids, so we much use ions (CaCl) to make the bacteria competent and able to accept foreign DNA. Start: 2 vials of competent bacteria (500 uL each) 9 Transformations U V (2 uL) dV (5 uL) dV + dI (5 uL) ddV (5 uL) ddV + ddI (5 uL) dV (10 uL) dV + dI (10 uL) ddV (10 uL) ddV + ddI (10 uL)

  1. Aliquot 5 uL and 10 uL of ligation reactions into eppendorfs
  2. Pipette 50 uL of competent DH5a
Personal tools