User:Sean P Corum/Notebook/PHIX174 Cell Free/2012/06/22: Difference between revisions

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===Characterization C: Expression of PHIX174 promoters fused to UTRX-deGFP.===
===Characterization C: Expression of PHIX174 promoters fused to UTRX-deGFP.===


* Yesterday, I digested PCR products with SphI and NcoI to create SphI-(PB-UTRB, PG-UTRG, PL-UTRL, and PL-PA-UTRA)-NcoI linkers.  Today, I performed gel extraction on the digest products.
* Yesterday, I digested PCR products with SphI and NcoI to create SphI-(PB-UTRB, PG-UTRG, PL-UTRL, and PL-PA-UTRA)-NcoI linkers.  Today, I performed [http://openwetware.org/wiki/Corum:Gel_Purification gel extraction] on the digest products.
* I also hybridized oligonucleotides to create SphI-(PA-UTRA, PD-UTRD, PF-UTRF)-NcoI linkers (100μM, 10μM, 1μM, and 100nM serial dilutions).
* I also hybridized oligonucleotides to create SphI-(PA-UTRA, PD-UTRD, PF-UTRF)-NcoI linkers (100μM, 10μM, 1μM, and 100nM serial dilutions).
* The next step will be to ligate, isolation plasmid clones, and sequence.
* The next step will be to ligate, isolation plasmid clones, and sequence.

Revision as of 12:24, 25 June 2012

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June 22, 2012

  • SC 15:43, 23 June 2012 (EDT):

Characterization B: Expression of PHIX174 promoters fused to UTR1-deGFP.

  • Performed miniprep plasmid isolation on the five clones.
  • Submitted for sequencing.

Characterization C: Expression of PHIX174 promoters fused to UTRX-deGFP.

  • Yesterday, I digested PCR products with SphI and NcoI to create SphI-(PB-UTRB, PG-UTRG, PL-UTRL, and PL-PA-UTRA)-NcoI linkers. Today, I performed gel extraction on the digest products.
  • I also hybridized oligonucleotides to create SphI-(PA-UTRA, PD-UTRD, PF-UTRF)-NcoI linkers (100μM, 10μM, 1μM, and 100nM serial dilutions).
  • The next step will be to ligate, isolation plasmid clones, and sequence.

Hypothesis 2: Gene L is necessary for phage propagation.

  • Need to design new primers (regular and mutagenic) for amplifying PHIX174 genome.


Notes

  • None.


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