IGEM:MIT/2007/Notebook/2007-8-1: Difference between revisions

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#Decide on constitutive promoter for CPX
#Decide on constitutive promoter for CPX
#*Transform promoter into DH5a
#*Transform promoter into DH5a
#*Plate transformants
#Check sequencing results for Mer/GFP 3A
#Check sequencing results for Mer/GFP 3A
#Perform flourescence assay on mer-gfp construct
#Perform flourescence assay on mer-gfp construct
 
#Clean out fridge


===Flourescence Assay Protocol===
===Flourescence Assay Protocol===
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# Repeat 4
# Repeat 4
# Put 150ul in plate reader
# Put 150ul in plate reader
<gallery>
Image:8-1-AHL-5Ai.jpg|AHL 5Ai
Image:8-1-AHL-5Aii.jpg|AHL 5Aii
Image:8-1-AHL-5Bi.jpg|AHL 5Bi
Image:8-1-AHL-5Bii.jpg|AHL 5Bii
Image:8-1-AHL-6Ai.jpg|AHL 6Ai
Image:8-1-AHL-6Aii.jpg|AHL 6Aii
Image:8-1-AHL-6Bi.jpg|AHL 6Bi
Image:8-1-AHL-7Ai.jpg|AHL 7Ai
Image:8-1-AHL-7Aii.jpg|AHL 7Aii
Image:8-1-AHL-7Aiii.jpg|AHL 7Aiii
Image:8-1-AHL-7Bi.jpg|AHL 7Bi
Image:8-1-AHL-7Bii.jpg|AHL 7Bii
Image:8-1-AHL-8Ai.jpg|AHL 8Ai
Image:8-1-AHL-8Aii.jpg|AHL 8Aii
Image:8-1-AHL-8Bi.jpg|AHL 8Bi
Image:8-1-AHL-8Bii.jpg|AHL 8Bii
Image:8-1-controlAi.jpg|Control Ai
Image:8-1-controlAii.jpg|Control Aii
Image:8-1-controlBi.jpg|Control Bi
Image:8-1-controlBii.jpg|Control Bii
</gallery>


===Sequencing Results===
===Sequencing Results===
*mer.I13500 in pSB1AT3 came back really crappy.  Biobrick prefix and suffix are partially present, cannot locate the sequence for GFP.
*mer.I13500 in pSB1AT3 came back really crappy.  Biobrick prefix and suffix are partially present, cannot locate the sequence for GFP.
*Need to send in the parts for sequencing.
*Need to send in the parts for sequencing.
*Will run a colony PCR as a diagnostic.
*Will run a colony PCR as a diagnostic?
 
===Constitutive Promoter===
Construct containing CPX and the polystyrene-binding peptide that will hopefully go into final system:
 
I14032/R0051.B0034.CPX-insert.
 
I14032 is a lacI repressable constitutive promoter.  R0051 is a lambda cl repressable promoter.
===Things That Got Cleaned Out===
Ask Semmie.

Latest revision as of 14:24, 2 August 2007

Agenda

  1. Decide on constitutive promoter for CPX
    • Transform promoter into DH5a
    • Plate transformants
  2. Check sequencing results for Mer/GFP 3A
  3. Perform flourescence assay on mer-gfp construct
  4. Clean out fridge

Flourescence Assay Protocol

  1. Draw sample of 1.5 ml from incubated cells with Hg
  2. Plot OD's
  3. Spin down 3 mins at 13000 rpm
  4. Aspirate and resuspend in 1.5 ml .9% NaCl
  5. Repeat 3
  6. Repeat 4
  7. Put 150ul in plate reader

Sequencing Results

  • mer.I13500 in pSB1AT3 came back really crappy. Biobrick prefix and suffix are partially present, cannot locate the sequence for GFP.
  • Need to send in the parts for sequencing.
  • Will run a colony PCR as a diagnostic?

Constitutive Promoter

Construct containing CPX and the polystyrene-binding peptide that will hopefully go into final system:

I14032/R0051.B0034.CPX-insert.

I14032 is a lacI repressable constitutive promoter. R0051 is a lambda cl repressable promoter.

Things That Got Cleaned Out

Ask Semmie.

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