Team:Hawaii/Notebook/2008-08-15

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(New page: {{Team:Hawaii/Header}} = Things we did today = == Wetlab work == ===Reconstruction of BB-pRl1383a=== :<strong> Grace</strong> :* PCR amplification of J33207 ::* ExoSAP'd PCR product :* R...)
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= Things we did today =
= Things we did today =
== Wetlab work ==
== Wetlab work ==
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===Reconstruction of BB-pRl1383a===
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===Reconstruction of BB-pRL1383a===
 +
[[Image:081508REdigest.jpg|right|thumb|200px|EtBr stained 1% agarose gel ran at 72V for 1.5 hours. Thirty-five microliters of RE digested BB-pRL1383a and fifty microliters of RE digested J33207 were loaded.]]
:<strong> Grace</strong>
:<strong> Grace</strong>
:* PCR amplification of J33207
:* PCR amplification of J33207
::* ExoSAP'd PCR product
::* ExoSAP'd PCR product
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:* RE digested of J33207 and BB-pRL1383a (w/ GFP)
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:* RE digested of J33207 and BB-pRL1383a (w/ GFP) with EcoRI and PstI
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:* Ran digests on a 0.8% agarose gel
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:* Ran digests on a 1% agarose gel at 72V for 1.5 hours.
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:* Extracted from bands from gel
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::* Ladder did not run well. Bands not resolving well.
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:* Ligated J33207 w/ BB-pRL1383a
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:* Redid PCR and RE digests with XbaI and PstI
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:* Transformed into DH5&alpha;
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== Drylab Work ==
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===Made 20X X-gal stock solution (20 mg/ml)===
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:<strong>Grace</strong>
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===Name of Task===
 
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:<strong> name of person/people who performed the task</strong>
 
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:* Summary of task and what was done. Link to experiment for detailed notes if necessary.
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===Restriction Digest===
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:* e.g. read through papers, worked on proposal, etc.
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:<strong>Margaret</strong>
 +
:*Digested J33207 (contains lacZ for blue/white screening), oriT--> these will be ligated so i can test the origin of transfer in a conjugation experiment.
 +
:*Digested I14032 and the aadA(BB) which was inserted into B0030 so I can ligate these two together.
 +
===Streak to verify antibiotic resistance===
 +
:<strong>Margaret</strong>
 +
 +
:*On an LB plate containing Sm&Sp, the omega ligation (from 8-12). This will be a verification that the omega ligation worked. As a positive control I streaked pSMC121 which contains the omega interposon, and as a negative control I streaked pSB1A2 (in case the plates have lost their effectiveness).
 +
 +
== Drylab Work ==
 +
 +
===Editing team website===
 +
:<strong> Grace</strong>
= Discussion =
= Discussion =
:*DH5&alpha; does not carry ''lacI<sup>q</sup>''; therefore, IPTG is not neccessary to induce the ''lac'' promoter.
:*DH5&alpha; does not carry ''lacI<sup>q</sup>''; therefore, IPTG is not neccessary to induce the ''lac'' promoter.
 +
:* IPTG is necessary for induction in DB3.1 cells.
 +
= Quote of the Day =
= Quote of the Day =
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<blockquote>''History is the only laboratory we have in which to test the consequences of thought.'' - Étienne Gilson</blockquote>
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<blockquote>''I'm waiting for the terminator to come in...'' - Krystle</blockquote>
{{Team:Hawaii/Footer}}
{{Team:Hawaii/Footer}}
__NOTOC__
__NOTOC__

Latest revision as of 03:16, 16 August 2008

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Things we did today

Wetlab work

Reconstruction of BB-pRL1383a

EtBr stained 1% agarose gel ran at 72V for 1.5 hours. Thirty-five microliters of RE digested BB-pRL1383a and fifty microliters of RE digested J33207 were loaded.
Grace
  • PCR amplification of J33207
  • ExoSAP'd PCR product
  • RE digested of J33207 and BB-pRL1383a (w/ GFP) with EcoRI and PstI
  • Ran digests on a 1% agarose gel at 72V for 1.5 hours.
  • Ladder did not run well. Bands not resolving well.
  • Redid PCR and RE digests with XbaI and PstI

Made 20X X-gal stock solution (20 mg/ml)

Grace


Restriction Digest

Margaret
  • Digested J33207 (contains lacZ for blue/white screening), oriT--> these will be ligated so i can test the origin of transfer in a conjugation experiment.
  • Digested I14032 and the aadA(BB) which was inserted into B0030 so I can ligate these two together.

Streak to verify antibiotic resistance

Margaret
  • On an LB plate containing Sm&Sp, the omega ligation (from 8-12). This will be a verification that the omega ligation worked. As a positive control I streaked pSMC121 which contains the omega interposon, and as a negative control I streaked pSB1A2 (in case the plates have lost their effectiveness).

Drylab Work

Editing team website

Grace

Discussion

  • DH5α does not carry lacIq; therefore, IPTG is not neccessary to induce the lac promoter.
  • IPTG is necessary for induction in DB3.1 cells.

Quote of the Day

I'm waiting for the terminator to come in... - Krystle


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