Team:Brown/Notebook/July18

From 2010.igem.org

Sunday, July 18 2010

Results of 7/17 test of competent cells:

XL1B

With 100 pg of RFP plasmid: 16 colonies
With 1 ng of RFP plasmid: ~150 colonies
With 10 ng of RFP plasmid: >800 colonies

XL1B

With 100 pg of empty pNoTat: ~50 colonies
With 1 ng of empty pNoTat: ~180 colonies
With 10 ng of empty pNoTat: a lot of colonies (thousands).

Preparation of Tat BioBricks from registry:

pAraC = R0080

Plate 1: 12E

AraC = C0080

Plate 1: 14L
Kan resistance

LacI = I732100

Plate 2: 10E

Terter = B0015

Plate 1: 23L

Punched hole, added 10 µl of ddH2O.

Used 3 mL of rehydrated DNA to transform XL1.

Plated and incubate at 37°C overnight.

7:00 PM: Removed 8 aliquots of competent cells (Xl1B) from yesterday and let them thaw on ice.

7:04 PM: Removed 3 null plates, 3 amp plates, 3 kan plates, and 3 tet plates from the fridge.

7:22 PM: Diluted BBa_K19005 (150 ng/µl) to 149 µl dH2O, results in 1 ng/µl concentration DNA.

Also, diluted Sosnick vector TrpR by diluting 1 µl of plasmid at 393 ng/µl into 392 µl H2O for a final concentration of 1 ng/µl.

Tat Parts List

pAraC__Bba_R0080__’10 Plate 1__12E__pSB1A2
RBS-ChFP-Terter__Bba_J06702__’10 Plate 2__8E__pSB1A2
AraC__Bba_C0080__’10 Plate 1__14L__pSB2K3
LacI__Bba_I732100__’10 Plate 2__10E__pSB1A3
Terter__Bba_B0015__’10 Plate 1__23L__pSB1AK3
pLac-RBS-CFP-Terter__Bba_J04421__’08 Plate 1__4G__pSB1A2

Accidentally diluted stock of BBa_K19005 with 149 µl dH2O, re-nanodropped and got a reading of 18.4 ng/µl. Added 1 µl of this to 17.4µl dH2O to make 1 ng/µl stock.

8:07 PM: Added 10 ng DNA to each aliquot and incubated for 20 minutes.

Later: Finished cell transformation protocol, placed in incubator.