Team:Hawaii/Notebook/2008-10-21

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== Wetlab work ==
== Wetlab work ==
===Insertion of lac-GFP device into BB-pRL1383a===
===Insertion of lac-GFP device into BB-pRL1383a===
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[[Image:102108REdigests.png|right|thumb|150px|EtBr stained 0.8% agarose gel ran at 60V for 90 minutes. Thirty microliters of the RE digest reaction were loaded into each well.]][[Image:102108J04430.png|right|thumb|150px|EtBr stained 0.8% agarose gel ran at 60V for 90 minutes. Ten microliters of PCR product were loaded.]]
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:<strong>  Grace</strong>
:<strong>  Grace</strong>
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[[Image:102108REdigests.png|right|thumb|EtBr stained 0.8% agarose gel ran at 60V for 90 minutes. Thirty microliters of the RE digest reaction were loaded into each well.]]
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:*Ran RE digests on gel
:*Ran RE digests on gel
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::* [[quantification of bands]]
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::*Bands were all incorrect size
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:* Extracted bands from gel
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:* Reinoculated TB+sp<sub>100</sub> for plasmid prep tomorrow
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:* Ligated J04430 and BB-pRL1383a
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:* PCR of J04430 from filter paper to verify part; used H/B primers
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:* Used 5&mu;l to transform DH5&alpha;
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===Antibiotic test for BB-pRL1383a===
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===[[Team:Hawaii/Antibiotic test for BB-pRL1383a]]===
:<strong> Grace</strong>
:<strong> Grace</strong>
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:* Made LB+sp<sub>variable</sub> plates with 40 &mu;l 20X X-gal
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:* Made LB+sp<sub>variable</sub> plates with 50 &mu;l 20X X-gal
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:* Transformed 100 &mu;l DH5&alpha; with 2 &mul; BB-pRL1383a (4 ng)
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:* Transformed 100 &mu;l DH5&alpha; with 2 &mu;l BB-pRL1383a (4 ng)
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:* Plated 50 &m;ul cells + SOB on each plate
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:* Plated 50 &mu;l cells + SOB on each LB+sp plate; plated 20 &mu;l untransformed cells + SOB on each control plate
:* Incubated at 37C for 2 days
:* Incubated at 37C for 2 days
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 +
===Construction of GFP secretion device===
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:<strong>Krystle</strong>
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:* Gel purified overnight restriction digested nir+rbs
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::* Loaded all of restriction digest into a 2% gel, which appeared blank after running for 1.5 hours
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:* Re-PCR nir+rbs and slr+gfpf from plasmid prep
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:* Restriction digest
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::*nir+rbs with EcoRI and SpeI
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::*slr+gfpf with XbaI and PstI
= Discussion =
= Discussion =

Latest revision as of 01:05, 30 October 2008

Projects Events Resources
Sponsors Experiments Milestones Protocols
Notebook (t) Meetings (t)

Things we did today

Wetlab work

Insertion of lac-GFP device into BB-pRL1383a

EtBr stained 0.8% agarose gel ran at 60V for 90 minutes. Thirty microliters of the RE digest reaction were loaded into each well.
EtBr stained 0.8% agarose gel ran at 60V for 90 minutes. Ten microliters of PCR product were loaded.
Grace
  • Ran RE digests on gel
  • Bands were all incorrect size
  • Reinoculated TB+sp100 for plasmid prep tomorrow
  • PCR of J04430 from filter paper to verify part; used H/B primers

Team:Hawaii/Antibiotic test for BB-pRL1383a

Grace
  • Made LB+spvariable plates with 50 μl 20X X-gal
  • Transformed 100 μl DH5α with 2 μl BB-pRL1383a (4 ng)
  • Plated 50 μl cells + SOB on each LB+sp plate; plated 20 μl untransformed cells + SOB on each control plate
  • Incubated at 37C for 2 days

Construction of GFP secretion device

Krystle
  • Gel purified overnight restriction digested nir+rbs
  • Loaded all of restriction digest into a 2% gel, which appeared blank after running for 1.5 hours
  • Re-PCR nir+rbs and slr+gfpf from plasmid prep
  • Restriction digest
  • nir+rbs with EcoRI and SpeI
  • slr+gfpf with XbaI and PstI

Discussion

Quote of the Day

History is the only laboratory we have in which to test the consequences of thought. - Étienne Gilson


[http://manoa.hawaii.edu/ Sponsor_UHM.gif][http://manoa.hawaii.edu/ovcrge/ Sponsor_OVCRGE.gif][http://www.ctahr.hawaii.edu Sponsor_CTAHR.gif]


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