Team:UI-Indonesia/September

From 2013.igem.org

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<h1>September 1st</h1>
 +
<li>Purification of PCR omega fragment</li>
 +
<li>Making gene ruler </li>
 +
<li>Running the purified product </li>
 +
<li>Nanodrop the purified product : 110 ng/μL</li>
 +
<li>Inoculating bacteria with pBluescript KS and pSB1C3 containg alpha fragment</li>
 +
<li>Spreading BL21 and Top10 on agar plates with chloramphenicol antibiotic to check the optimum chloramphenicol concentration.</li>
 +
 +
<h1>September 3rd</h1>
 +
<li>Nanodrop the large scale isolation product:
 +
<ul>pBluesript KS: 69.5 ng/μL</ul>
 +
<ul>Alpha fragment : 179.5 ng/μL</ul></li>
 +
<li>Restricting pBluescript KS using EcoRv</li>
 +
<li>Nanodrop the restricted plasmid</li>
 +
<li>Making competent cells</li>
 +
<h1>September 4th</h1>
 +
<li>Testing te chloramphenicol concentration</li>
 +
<li>Testing the enzymatic activity from PstI</li>
 +
<li>Making agar plates with different concentration of chloramphenicol</li>
 +
<li>Running the result of enzymatic activity test</li>
 +
 +
<h1>September 5th</h1>
 +
<li>Re-testing the enzymatic activity using different buffers</li>
 +
<li>Running</li>
 +
<li>Checking the chloramphenicol test result</li>
 +
<li>Transforming:
 +
<ul>Omeg fragment-pBluescript KS</ul>
 +
<ul>(G4S)4 -pSB1C3 (ligated 3/9)</ul>
 +
<ul>Lox511-psB1C3 (ligated 3/9)</ul>
 +
<ul>Lox511-psB1C3 (ligated 25/8)</ul>
 +
<ul>Lox511-psB1C3 (ligated 25/8)</ul>
 +
<ul>(G4S)4 -pSB1C3 (ligated 25/8)</ul>
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<ul>pQE80L</ul>
 +
<ul>Re-ligating omega  fragment-pSB1C3</ul></li>
 +
 +
<h1>September 6th</h1>
 +
<li>Restricting:
 +
<ul>Omega fragment + EcoRI</ul>
 +
<ul>pSB1C3</ul>
 +
<ul>Omega fragmetn +XbaI</ul>
 +
<ul>pBluscript KS + XbaI</ul>
 +
<ul>Alpha fragment + XbaI</ul></li>
 +
<li>Running</li>
 +
<li>Nanodrop the purified digested plasmid
 +
<ul>Omega fragment + EcoRI = 7.2 ng/μL</ul>
 +
<ul>pSB1C3 = 3.8 ng/μL</ul>
 +
<ul>Omega fragment + XbaI = 7.7 ng/μL</ul>
 +
<ul>pBluescript KS + XbaI = 9.8 ng/μL</ul>
 +
<ul>Alpha fragment + Xba I =  0.35 ng/μL</ul></li>
 +
        <li>Running</li>
 +
        <li>Inoculating bacteria with pSB1C3-alpha fragment in 100mL broth</li>
 +
 +
<h1>September 9th</h1>
 +
<li>Large scale pSB1C3-alpha fragment isolation</li>
 +
<li>Restricting pBluescript KS using EcoRV</li>
 +
<li>Digesting linearized pSB1C3 and linearized omega fragment using PstI</li>
 +
<li>Making 750mL LB Broth</li>
 +
<li>Making agar plates</li>
 +
<li>Digesting pSB1C3-alpha fragment</li>
 +
<li>Purifying the digested using LMA</li>
 +
<li>Blunting Lox511 and (G4S)4 linker</li>
 +
<li>Running</li>
 +
 +
<h1>September 11th</h1>
 +
<li>Small scale isolation</li>
 +
<li>Running the small scale isolation product</li>
 +
 +
<h1>September 13th</h1>
 +
<li>Re-run the small scale iolation product</li>
 +
<li>Inoculate the bacteria containing pSB1C3=omega fragment</li>
 +
 +
<h1>September 14th</h1>
 +
<li>Running small scale isolation product</li>
 +
<li>PCR Colony</li>
 +
<li>Large scale isolation</li>
 +
<li>Re-ligating omega fragment with pSB1C3</li>
 +
 +
<h1>September 15th</h1>
 +
<li>Running
 +
<ul>Digested pQE+PstI</ul>
 +
<ul>PQE</ul>
 +
<ul>Digested alpha fragment+PstI</ul>
 +
<ul>Alpha fragment</ul>
 +
<ul>Marker</ul>
 +
<ul>5</ul>
 +
<ul>8</ul>
 +
<ul>9</ul>
 +
<ul>15</ul>
 +
<ul>Large scale isolation pBluescript KS-omega</ul></li>
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{{:Team:UI-Indonesia/footer}}
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Revision as of 04:09, 28 September 2013




September

September 1st

  • Purification of PCR omega fragment
  • Making gene ruler
  • Running the purified product
  • Nanodrop the purified product : 110 ng/μL
  • Inoculating bacteria with pBluescript KS and pSB1C3 containg alpha fragment
  • Spreading BL21 and Top10 on agar plates with chloramphenicol antibiotic to check the optimum chloramphenicol concentration.

    • September 3rd

  • Nanodrop the large scale isolation product:
      pBluesript KS: 69.5 ng/μL
      Alpha fragment : 179.5 ng/μL
  • Restricting pBluescript KS using EcoRv
  • Nanodrop the restricted plasmid
  • Making competent cells

    • September 4th

  • Testing te chloramphenicol concentration
  • Testing the enzymatic activity from PstI
  • Making agar plates with different concentration of chloramphenicol
  • Running the result of enzymatic activity test

    • September 5th

  • Re-testing the enzymatic activity using different buffers
  • Running
  • Checking the chloramphenicol test result
  • Transforming:
      Omeg fragment-pBluescript KS
      (G4S)4 -pSB1C3 (ligated 3/9)
      Lox511-psB1C3 (ligated 3/9)
      Lox511-psB1C3 (ligated 25/8)
      Lox511-psB1C3 (ligated 25/8)
      (G4S)4 -pSB1C3 (ligated 25/8)
      pQE80L
      Re-ligating omega fragment-pSB1C3

    • September 6th

  • Restricting:
      Omega fragment + EcoRI
      pSB1C3
      Omega fragmetn +XbaI
      pBluscript KS + XbaI
      Alpha fragment + XbaI
  • Running
  • Nanodrop the purified digested plasmid
      Omega fragment + EcoRI = 7.2 ng/μL
      pSB1C3 = 3.8 ng/μL
      Omega fragment + XbaI = 7.7 ng/μL
      pBluescript KS + XbaI = 9.8 ng/μL
      Alpha fragment + Xba I = 0.35 ng/μL
  • Running
  • Inoculating bacteria with pSB1C3-alpha fragment in 100mL broth

    • September 9th

  • Large scale pSB1C3-alpha fragment isolation
  • Restricting pBluescript KS using EcoRV
  • Digesting linearized pSB1C3 and linearized omega fragment using PstI
  • Making 750mL LB Broth
  • Making agar plates
  • Digesting pSB1C3-alpha fragment
  • Purifying the digested using LMA
  • Blunting Lox511 and (G4S)4 linker
  • Running

    • September 11th

  • Small scale isolation
  • Running the small scale isolation product

    • September 13th

  • Re-run the small scale iolation product
  • Inoculate the bacteria containing pSB1C3=omega fragment

    • September 14th

  • Running small scale isolation product
  • PCR Colony
  • Large scale isolation
  • Re-ligating omega fragment with pSB1C3

    • September 15th

  • Running
      Digested pQE+PstI
      PQE
      Digested alpha fragment+PstI
      Alpha fragment
      Marker
      5
      8
      9
      15
      Large scale isolation pBluescript KS-omega

    • Retrieved from "http://2013.igem.org/Team:UI-Indonesia/September"