Latest revision as of 08:44, 15 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.

Margaret

5ml ON culture, LB with Amp100 for: colonies with the correct insert from a transformation completed on 9/7.

Construct	# colonies checked	correct (these are plasmid prepped)
pSB1A3::plac+RBS(B0030)+rep	8	0
pSB1A3::plac+RBSB0034+aadA(prl1383a)colony 1	8	2, 6
pSB1A3::plac+RBSB0034+aadA(BB)colony9	8	2, 3, 4, 5, 6, 8

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

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

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 ===Verification of transformants===
 :<strong> Grace</strong>
-[[Image:091208colonyPCR.jpg|right|thumb|400px|EtBr stained 2% agarose gel ran at 95V for 1 hour. Two microliters of PCR product were loaded into each well.]]
+[[Image:091208colonyPCR.png|right|thumb|400px|EtBr stained 2% agarose gel ran at 95V for 1 hour. Two microliters of PCR product were loaded into each well.]]
 {|class=wikitable border=1 align=center
 !Construct