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

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).
 * [[Image:Ligation_miniprep_gels.doc|Click here to see Gels]]

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