Haynes Lab:Notebook/Characterizing AHL quorum sensing homologs/2013/10/28

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==mm/dd/yyyy==
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==10/28/2013==
Miniprepped J04450 that I started growing on Saturday. They were in the incubator for a long time and were super red. <br>
Miniprepped J04450 that I started growing on Saturday. They were in the incubator for a long time and were super red. <br>
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[[image:Pretty mRFP liq cultures.JPG|270px|Pretty mRFP]]
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<br><br>
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'''Miniprep'''<br>
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{| {{table}}
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|-
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|260 ||0.034
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|-
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|280 ||0.021
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|-
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|260/280|| 1.631
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|-
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|ng/µL ||33.893
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|-
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|}
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<br>
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'''Restriction digest'''<br>
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cut 15ul of 33ng/ul, so about 0.5ug of plasmid. Cut with E/X in a 20ul rxn for 25 minutes at 37°C<br>
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<br>
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Gel of cut J04450 [[image:13.10.28_J04450.png‎|J04450 E/X]]
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<br>Cut out and put the gel slice at -20°C<br><br>
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Gel image
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13.10.28_J04450.png‎
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PCR image
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13.10.28_PCR_cropped.png‎
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PCR ''need to edit this to include the next lane. lane 1 is P087/P088 and lane 2 is P089/P090, both I13522'' [[image:13.10.28_PCR_cropped.png‎|PCR]]
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<br><br>
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Ran another PCR reaction to text which primer wasn't working for the sender backbone. Used the two sender backbone primers, P089 and P090 with the matching F or R primer from the pLux vectors primers which were shown to work. Rxn 3 is positive control.<br>
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1
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:15ul reaction
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:P087/P090
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2
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:15ul reaction
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:P089/P088
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3
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:10ul
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:P087/P088
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10/28/2013

Miniprepped J04450 that I started growing on Saturday. They were in the incubator for a long time and were super red.

Pretty mRFP



Miniprep

260 0.034
280 0.021
260/280 1.631
ng/µL 33.893

Restriction digest
cut 15ul of 33ng/ul, so about 0.5ug of plasmid. Cut with E/X in a 20ul rxn for 25 minutes at 37°C

Gel of cut J04450 J04450 E/X
Cut out and put the gel slice at -20°C

PCR need to edit this to include the next lane. lane 1 is P087/P088 and lane 2 is P089/P090, both I13522 PCR

Ran another PCR reaction to text which primer wasn't working for the sender backbone. Used the two sender backbone primers, P089 and P090 with the matching F or R primer from the pLux vectors primers which were shown to work. Rxn 3 is positive control.
1

15ul reaction
P087/P090

2

15ul reaction
P089/P088

3

10ul
P087/P088


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