Revision as of 05:10, 13 September 2008

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Things we did today

Grace

EtBr stained 2% agarose gel ran at 95V for 1 hour. Two microliters of PCR product were loaded into each well.

Margaret, Krystle, &Grace

5ml culture grown over-night in TB (terrific broth), in the morning 125ml SOB was inoculated with 12.5mL(1), and 2 flasks with 50ml (2&3) SOB were inoculated with 5ml of the ON culture.

The cells were spun down at 2147 x g, 4°C, 30 minutes. The yield was low (1/4 of the tip of a 50ml centrifuge tube), so the culturing was repeated, except that the starting OD was 0.14 and the final OD was more than 0.3 but less than 0.6 (with the same amount of cells yielded).
The cells are being cultured again tonight to try again tomorrow. IF we get the same amount of cells, we will continue with the competent cell making procedure.

Wait, you have an internet boyfriend? - KS, to Norman

[http://manoa.hawaii.edu/ ][http://manoa.hawaii.edu/ovcrge/ ][http://www.ctahr.hawaii.edu ]

@@ Line 65: / Line 65: @@
 |}
-:*Colony PCR of 3A, plac+rbs transformants
+===Competent Cells===
+:<strong> Margaret, Krystle, &Grace</strong>
+:*Several attempts were made to make more DH5-alpha competent cells.
+::*5ml culture grown over-night in TB (terrific broth), in the morning 125ml SOB was inoculated with 12.5mL(1), and 2 flasks with 50ml (2&3) SOB were inoculated with 5ml of the ON culture.
+{|class=wikitable border=1
+!Flask #
+!OD550 (1hr)
+!OD550 (1.5hr)
+|-
+|1
+|0.2056
+|0.3432
+|-
+|2
+|0.2006
+|0.3312
+|-
+|3
+|0.2547
+|0.4184
+|}
+:*The cells were spun down at 2147 x g, 4&deg;C, 30 minutes. The yield was low (1/4 of the tip of a 50ml centrifuge tube), so the culturing was repeated,  except that the starting OD was 0.14 and the final OD was more than 0.3 but less than 0.6 (with the same amount of cells yielded).
+:*The cells are being cultured again tonight to try again tomorrow. IF we get the same amount of cells, we will continue with the competent cell making procedure.