IGEM:IMPERIAL/2006/LabCalendar/2006-8-1

• IGEM:IMPERIAL/2006/LabCalendar/2006-8-1 Events
• Day in the lab
• Project Progress

Mini-prep parts

• 7A->3O not successful first mini-prep, Dr. Mann suggested redoing ligation. However, upon second attempt to mini-prep, one colony appears to have successfully taken up the ligated vector (Gel E, Lane 2)
• 12D->2H unsucessful ligation according to miniprep results (Gel B and Gel C lanes 2 and 3). Need to religate tomorrow.
• 12D->24A not successful first mini-prep, redoing miniprep this afternoon. Successful mini-prep results can be seen in Gel C (Lanes 4 and 5) and Gel D (Lanes 2 and 3).
  • File:Ligation miniprep gels.doc

Re-ligation of 7A->3O

• Digests
  • 7A with X & P, using yellow buffer
  • 3O with S & P, using orange buffer
• Run on large gel
  • Successful...was able to cut out insert and vector
  • 7A extracted 756 bp (insert)
  • 3O extracted 2091 bp (vector)
• Preform ligation after gene cleaning
  • 16 uL insert
  • 1 uL vector
  • 2 uL ligase buffer
  • 1 uL ligase

M9 broth

• We want a total of 200 mL
  • 100 mL 2x M9 salts
  • 6 mL 10 mg/mL thiamine
  • 2 mL 40% glycerol
  • 4 mL 10% casamino acids
  • 4 mL 0.1 M MgSO4
  • 40 uL 0.5M CaCl2
  • Make up to 200 mL with sterile water
• See Endy Lab Protocol on OWW

Electroporations Tomorrow

• 7A->3O ligation
• Part I13521 well 18I, plate 1

Maxi-prep tomorrow

• 12D->2H
• J04500, 16P
• C0060, 5M
• B0015, 1I
• Recultured bacteria in 100 mL LB with Amp with 100 uL bacteria culture
• Set for at least 14 hours in 37C shaker