Team:Wash U/Calendar-Home/15 July 2009

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Latest revision as of 21:44, 15 July 2009

Wednesday July 15, 2009

-Gel purification from yesterday yielded low DNA concentrations

-Started anew today with same digestions (OmpC, GFP) but only gel purifying the destination plasmids

Gel layout:

1) 100 bp ladder

2) OmpC + vector (2187 bp)

3) OmpC + vector

4) GFP (876 bp)

5) GFP

6) Destination plasmid, pSB1K3 (2206 bp)

7) pSB1K3

8) 1 kb ladder

Gel #2:

1) 1 kb ladder

2) OmpC + vector (2187 bp)

3) same

4) same

5) same

6) GFP (876 bp)

7) same

Note: This second digestion was run using the GFP sample from the 7-1 miniprep and the OmpC sample from the 7-2 miniprep. The 7-2 OmpC DNA was very dilute and required about 74 microliters to obtain 700 ng, so enzyme and buffer volumes were adjusted for a total reaction volume of 100 microliters (requiring four gel lanes).

Ligation reactions:

1: 6 microliters OmpC, 8 microliters GFP, 4 microliters pSB1K3, 2 microliters buffer, 1 microliter enzyme

2: 4 microliters OmpC, 6 microliters GFP, 4 microliters pSB1K3, 2 microliters buffer, 1 microliter enzyme, 3 microliters water

-Team meeting #6

-Made new Kanamycin (10ug/ml) LB plates

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@@ Line 6: / Line 6: @@
 -Gel purification from yesterday yielded low DNA concentrations
--Started anew today with same digestions but only gel purifying the destination plasmids
+-Started anew today with same digestions (OmpC, GFP) but only gel purifying the destination plasmids
--Team meeting
+Gel layout:
+) 100 bp ladder
+) OmpC + vector (2187 bp)
+) OmpC + vector
+) GFP (876 bp)
+) GFP
+) Destination plasmid, pSB1K3 (2206 bp)
+) pSB1K3
+) 1 kb ladder
+Gel #2:
+) 1 kb ladder
+) OmpC + vector (2187 bp)
+) same
+) same
+) same
+) GFP (876 bp)
+) same
+Note: This second digestion was run using the GFP sample from the 7-1 miniprep and the OmpC sample from the 7-2 miniprep.  The 7-2 OmpC DNA was very dilute and required about 74 microliters to obtain 700 ng, so enzyme and buffer volumes were adjusted for a total reaction volume of 100 microliters (requiring four gel lanes).
+Ligation reactions:
+: 6 microliters OmpC, 8 microliters GFP, 4 microliters pSB1K3, 2 microliters buffer, 1 microliter enzyme
+: 4 microliters OmpC, 6 microliters GFP, 4 microliters pSB1K3, 2 microliters buffer, 1 microliter enzyme, 3 microliters water
+-Team meeting #6
+-Made new Kanamycin (10ug/ml) LB plates