Team:NCTU Formosa/notes

From 2013.igem.org

(Difference between revisions)
Line 1,690: Line 1,690:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>1</p></div>
+
<div class="daybar"><p></p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
-
<li class="caldot green"></li>
 
-
<li class="caldot green"></li>
 
</ul>
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</div>
</div>
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>PCR of insert fragment [B0034+PBSII+ilvC] OK!</p></li>
 
-
<li class="green e2"><p>DNA Sequencing OK!</p></li>
 
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day brn">
<div class="day brn">
-
<div class="daybar"><p>2</p></div>
+
<div class="daybar"><p>1</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
</ul>
</ul>
</div>
</div>
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<div class="open">
<div class="open">
<ul>
<ul>
 +
<li class="green e6"><p>PCR and single colony isolation of TetR+B0015+PSB1K3</p></li>
 +
<li class="green e6"><p>Electrophoresis of PCR product made at this morning and Plac(5/30 Ar) OK</p></li>
 +
<li class="green e6"><p>Digestion:[Ter(B0015)]XP</p></li>
 +
<li class="green e6"><p>Transformation of Ter(B0015) on LB-A plate & mGFP on LB-K plate & 37oC RBS+mGFP+Ter(B0015) on LB-C plate</p></li>
 +
<li class="green e6"><p>Ligation: insert 37oC RBS[ES] & Ter(B0015)[XP]/ vector pSB1C3[EP]</p></li>
 +
<li class="green e6"><p>Single colony isolation from Ter(B0015) LB-A plate, and cultivation in liquid LB-A</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<!---------------------------------------- week start ---------------------------------------->
<!---------------------------------------- week start ---------------------------------------->
-
<div class="week mweek">
+
<div class="week bigwk">
<div class="day">
<div class="day">
-
<div class="daybar"><p>3</p></div>
+
<div class="daybar"><p>2</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
Line 1,732: Line 1,744:
<div class="open">
<div class="open">
-
                                                                <ul>
+
<ul>
-
<li class="green e2"><p>mini-prep of cultivated B0034+PBSII+ilvC E. coli &amp; pLac+B0034+Zif268+AlsS E. coli</p></li>
+
<li class="green e6"><p>Ligation: insert [TetR]ES+[B0015]XP/vector [PSB1K3]EP and transformation of this part.</p></li>
-
                                                                </ul>
+
<li class="green e6"><p>Ligation: B0032+ho1&B0034+ho1</p></li>
-
                                                                <ul>
+
<li class="green e6"><p>Transformation of B0032+ho1&B0034+ho1</p></li>
-
<li class="green e2"><p>mini-prep of cultivated HivC+ilvD-12,20 E. coli </p></li>
+
<li class="green e6"><p>Cultivation of pSB1C3(C20&C10)with liquid LB.</p></li>
-
                                                                </ul>
+
<li class="green e6"><p>PCR of insert fragment[37oC RBS+mGFP+Ter(B0015)]</p></li>
 +
<li class="green e6"><p>Transformation of mRFP on LB-A plate& LB-C plate& LB-K plate-Mini-prep of cultivated Ter(B0015) E.coli.</p></li>
 +
</ul>
</div>
</div>
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>4</p></div>
+
<div class="daybar"><p>3</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 1,751: Line 1,771:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>digestion : HivC+ilvD-12,20 [XP]</p></li>
+
<li class="green e7"><p>PCR of Plac &TetR+B0015</p></li>
 +
<li class="green e7"><p>Mini-prep of pSB1C3</p></li>
 +
<li class="green e7"><p>Cultivation of B0032+ho1&B0034+ho1 with LB-A plate PCR of B0032+ho1&B0034+ho1</p></li>
 +
<li class="green e7"><p>Electrophoresis of B0032+ho1&B0034+ho1──NOT OK.</p></li>
 +
<li class="green e7"><p>Single colony isolation from 37oC RBS+mGFP+Ter(B0015) LB-K plate, and cultivation in liquid LB-K & mRFP LB-A, and cultivation in liquid LB-K</p></li>
 +
<li class="green e7"><p>Mini-prep of cultivated 37oC RBS+mGFP+Ter(B0015) E.coli</p></li>
 +
<li class="green e7"><p>Digestion:[37oC RBS+mGFP+Ter(B0015)]XP</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>5</p></div>
+
<div class="daybar"><p>4</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>Design the primer for pLac+B0034+Zif268+AlsS point mutation (pm)</p></li>
+
<li class="green e8"><p>Electrophoresis of  Plac .TetR+B0015 OK</p></li>
 +
<li class="green e8"><p>Cultivation of Plac.TetR+B0015 in liquid LB-A tubes. PCR of B0032+ho1&B0034+ho1</p></li>
 +
<li class="green e8"><p>Electrophoresis of B0032+ho1(OK)&B0034+ho1(After PCR)</p></li>
 +
<li class="green e8"><p>Mini-prep of B0032+ho1. Digestion: [B0032+ho1]XP.</p></li>
 +
<li class="green e8"><p>Cultivation of ho1 with liquid LB-K</p></li>
 +
<li class="green e8"><p>Single colony isolation from mRFP LB-A plate, and cultivation in liquid LB-A</p></li>
 +
<li class="green e8"><p>Digestion:[37oC RBS+mGFP+Ter(B0015)]XP</p></li>
 +
<li class="green e8"><p>PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)].</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>6</p></div>
+
<div class="daybar"><p>5</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e4"><p>digestion : B0034 [ES] &amp; HivC [XP] &amp; pSB1C3 [EP] &amp; HivC [EP]</p></li>
+
<li class="green e8"><p>Mini-prep of cultivated Plac and TetR+B0015 E.coli and then conduct digestion :[Plac]P&[TetR+B0015]P</p></li>
-
<li class="green e4"><p>ligation : insert B0034 [ES] &amp; HivC [XP]/vector pSB1C3 [EP]</p></li>
+
<li class="green e8"><p>Electrophoresis of [Plac ]P and [TetR+B0015]P,[Plac]XP and [TetR+B0015]XP</p></li>
-
<li class="green e4"><p>ligation : insert HivC [EP]/vector pSB1C3 [EP]</p></li>
+
<li class="green e8"><p>Ligation: insert[PompC+B0034]ES+[LacI+J61048]XP/vector [PSB1A3]EP</p></li>
-
<li class="green e4"><p>transformation of B0034+ HivC &amp; HivC ,and cultivation on LB-C plate</p></li>
+
<li class="green e8"><p>Electrophoresis of B0032+ho1 mini&B0032+ho1(digested)&pSB1C3 mini&pSB1C3(digested)&ho1 mini&ho1(digested)</p></li>
 +
<li class="green e8"><p>Mini-prep of ho1</p></li>
 +
<li class="green e8"><p>Digestion: [ho1]XP</p></li>
 +
<li class="green e8"><p>Digestion:[37oC RBS+mGFP+Ter(B0015)]XP</p></li>
 +
<li class="green e8"><p>Transformation of mRFP on LB-A plate</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>7</p></div>
+
<div class="daybar"><p>6</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>PCR of insert fragment [B0034+ HivC] &amp; [HivC] OK</p></li>
+
<li class="green e7"><p>Transformation of PompC+B0034+LacI+J61048+PSB1A3</p></li>
 +
<li class="green e7"><p>Ligation: insert [PompC+B0034]ES+[LacI+J61048]XP/vector [PSB1A3]EP</p></li>
 +
<li class="green e7"><p>Electrophoresis of B0032+ho1 mini&B0032+ho1(digested)&ho1 mini&ho1 (digested).</p></li>
 +
<li class="green e7"><p>Single colony isolation from mRFP LB-A plate, and cultivation in liquid LB-A & LB-K & LB</p></li>
 +
<li class="green e7"><p>Digestion:[Ter(B0015)]EX</p></li>
 +
<li class="green e7"><p>Ligation: insert 37oC RBS+mGFP[ES]&Ter(B0015)[EX]</p></li>
 +
<li class="green e7"><p>Transformation of 37oCRBS+mGFP+Ter(B0015) on LB-K plate & LB-A plate, mRFP on LB-A plate</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>8</p></div>
+
<div class="daybar"><p>7</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>Design the primer for B0034+PBSII+ilvC point mutation (pm)</p></li>
+
<li class="green e4"><p>Transformation of PompC+B0034+LacI+J61048 again</p></li>
-
<li class="green e2"><p>Single colony isolation from HivC, B0034+HivC &amp; ilvD+37℃ RBS LB-C plate, and cultivation in liquid LB-C</p></li>
+
<li class="green e4"><p>Single colony isolation of TetR+B0015 Kr E.coli</p></li>
 +
<li class="green e4"><p>Digestion:[LacI+J61048]ES. PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)]</p></li>
 +
<li class="green e4"><p>Single colony isolation from 37oCRBS+mGFP+Ter(B0015) LB-A plate, and cultivation in liquid LB-A.</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day brn">
<div class="day brn">
-
<div class="daybar"><p>9</p></div>
+
<div class="daybar"><p>8</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e3"><p>mini-prep of cultivated HivC &amp; B0034+HivC &amp; ilvD+37℃ RBS E. coli </p></li>
+
<li class="green e8"><p>PCR + single colony isolation of PompC+B0034+LacI+J61048 E.coli</p></li>
-
<li class="green e3"><p>Testing the temperature of the point mutation between of HivC &amp; ilvD by the m.p 50℃ of PCR</p></li>
+
<li class="green e8"><p>Electrophoresis of PompC+B0034+LacI+J61048 OK</p></li>
-
<li class="green e3"><p>ligation: insert B0034+HivC [ES] &amp; ilvD [XP]/vector pSB1A3 [EP];insert B0034+HivC [ES] &amp; ilvD+37℃ RBS [XP]/vector pSB1A3 [EP]
+
<li class="green e8"><p>Ligation: insert [LacI+J61048]ES+[Plac]XP/vector[PSB1C3]EP</p></li>
-
</p></li>
+
<li class="green e8"><p>Transformation of LacI+J61048+Plac on LB-C plate</p></li>
 +
<li class="green e8"><p>Cultivation of PompC+B0034+LacI+J61048 in LB-A tubes & TetR+B0015 in LB-K tube & PSB1A3 in LB-A tube &PSB1C3 in LB-C tube.</p></li>
 +
<li class="green e8"><p>Mini-prep of cultivated 37oCRBS+mGFP+Ter(B0015) E.coli</p></li>
 +
<li class="green e8"><p>Digestion:[ 37oCRBS+mGFP+Ter(B0015)]XP</p></li>
 +
<li class="green e8"><p>PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)].</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<!---------------------------------------- week start ---------------------------------------->
<!---------------------------------------- week start ---------------------------------------->
-
<div class="week mweek">
+
<div class="week bigwk">
<div class="day">
<div class="day">
-
<div class="daybar"><p>10</p></div>
+
<div class="daybar"><p>9</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>transformation of B0034+ HivC+ ilvD &amp; B0034+ HivC+ ilvD+37℃ RBS ,and cultivation on LB-A plate</p></li>
+
<li class="green e4"><p>Mini-prep from cultivated PompC+B0034+LacI+J61048 Eco.li&PSB1C3 E.coli</p></li>
-
<li class="green e2"><p>Testing the temperature of the point mutation between of pLac+B0034+Zif268 &amp; AlsS by the m.p 55℃ of PCR failed</p></li>
+
<li class="green e4"><p>digestion : [PompC+B0034+LacI+J61048]ES&[PSB1C3]EP</p></li>
 +
<li class="green e4"><p>Electrophoresis of [PompC+B0034+LacI+J61048] ES& [LacI+J61048]ES&[PSB1C3]EP.</p></li>
 +
<li class="green e4"><p>Single colony isolation from 37oCRBS + mGFP+Ter(B0015)</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>11</p></div>
+
<div class="daybar"><p>10</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>DNA Sequencing B0034+HivC &amp; HivC OK</p></li>
+
<li class="green e3"><p>Mini-prep of cultivated 37oCRBS+mGFP+Ter(B0015)  E.coli</p></li>
-
<li class="green e2"><p>PCR of insert fragment [B0034+ HivC+ilvD] &amp; [B0034+ HivC+ilvD+37℃ RBS] OK</p></li>
+
<li class="green e3"><p>Digestion:[ 37oCRBS+mGFP+Ter(B0015)]XP</p></li>
 +
<li class="green e3"><p>Single colony isolation from 37oCRBS+mGFP+Ter(B0015) LB-A plate, and cultivation in liquid LB-A.</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>12</p></div>
+
<div class="daybar"><p>11</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
Line 1,892: Line 1,971:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>Single colony isolation from B0034+ HivC+ilvD+37℃ RBS &amp; B0034+ HivC+ ilvD LB-A plate, and cultivation in liquid LB-A</p></li>
+
<li class="green e2"><p>PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)]</p></li>
-
<li class="green e2"><p>Single colony isolation from HivC(TA) LB-A plate, and cultivation in liquid LB-A</p></li>
+
<li class="green e2"><p>Transformation of mRFP on LB-K plate.</p></li>
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>13</p></div>
+
<div class="daybar"><p>12</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
-
<li class="caldot green"></li>
 
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>mini-prep of cultivated B0034+ HivC+ilvD+37℃ RBS , B0034+ HivC+ ilvD &amp; HivC(TA) E. coli </p></li>
+
<li class="green"><p>Single colony isolation from mRFP LB-K plate, and cultivation in liquid LB-K</p></li>
-
<li class="green e2"><p>Testing the temperature of the point mutation between of pLac+B0034+Zif268 &amp; AlsS by the m.p 50 &amp; 52℃ of PCR----50℃ is better</p></li>
+
</ul>
</ul>
</div>
</div>
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</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>14</p></div>
+
<div class="daybar"><p>13</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 1,925: Line 2,006:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>PCR of insert fragment [pLac+B0034+Zif268+AlsS] with designed primer for point mutation -----NOT OK</p></li>
+
<li class="green e5"><p>Ligation: insert[PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1C3]EP and insert[TetR]ES+[B0015]XP/vector[PSB1C3]EP.</p></li>
 +
<li class="green e5"><p>Mini-prep of cultivated mRFP E.coli</p></li>
 +
<li class="green e5"><p>Digestion:[mRFP]ES & [Ter(J61048)]XP & [37OCRBS+mGFP]XP</p></li>
 +
<li class="green e5"><p>Ligation: insert 37oCRBS+LuxR[ES]&37oCRBS+mGFP[XP]  /vector pSB1A3[EP]</p></li>
 +
<li class="green e5"><p>Transformation of 37oCRBS +LuxR+37oCRBS+mGFP on LB-A plate & mRFP +Ter(J61048) on LB-C plate& Ter(J61048) on LB-A plate.</p></li>
</ul>
</ul>
</div>
</div>
Line 1,931: Line 2,016:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>15</p></div>
+
<div class="daybar"><p>14</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 1,940: Line 2,029:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>DNA Sequencing B0034+ HivC+ilvD+37℃ RBS &amp; B0034+ HivC+ ilvD OK</p></li>
+
<li class="green e5"><p>Transformation of  PompC+B0034+LacI+J61048+PSB1C3 and TetR+B0015+PSB1C3</p></li>
 +
<li class="green e5"><p>Cultivation of B0032+ho1 with liquid LB-A. PCR of insert fragment[Ter(B0015)]</p></li>
 +
<li class="green e5"><p>Single colony isolation from Ter(J61048) LB-A plate, and cultivation in liquid LB-A & 37OCRBS+mGFP on LB-K plate, and cultivation in liquid LB-K</p></li>
 +
<li class="green e5"><p>Transformation of 37oCRBS +LuxR+37oCRBS+mGFP on LB-A plate & mRFP +Ter(J61048) on LB-C plate</p></li>
 +
<li class="green e5"><p>Mini-prep of cultivated Ter (J61048)& 37oCRBS+mGFP E.coli</p></li>
</ul>
</ul>
</div>
</div>
Line 1,946: Line 2,039:
</div>
</div>
<div class="day brn">
<div class="day brn">
-
<div class="daybar"><p>16</p></div>
+
<div class="daybar"><p>15</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
</ul>
</ul>
</div>
</div>
Line 1,954: Line 2,051:
<div class="open">
<div class="open">
<ul>
<ul>
 +
<li class="green e4"><p>Digestion:[B0032]XP</p></li>
 +
<li class="green e4"><p>Transformation of  PompC+B0034+LacI+J61048 +PSB1C3and TetR+B0015 +PSB1C3 again</p></li>
 +
<li class="green e4"><p>Ligation: insert[PompC]ES+[B0032]XP/vector[PSB1K3]EP, then transformation of this part</p></li>
 +
<li class="green e4"><p>Mini-prep of B0032+ho1. PCR of insert fragment[37oCRBS + LuxR + 37oCRBS+mGFP]&[mRFP+Ter(J61048)].</p></li>
</ul>
</ul>
</div>
</div>
Line 1,961: Line 2,062:
<!---------------------------------------- week end ---------------------------------------->
<!---------------------------------------- week end ---------------------------------------->
<!---------------------------------------- week start ---------------------------------------->
<!---------------------------------------- week start ---------------------------------------->
-
<div class="week">
+
<div class="week bigwk">
<div class="day">
<div class="day">
-
<div class="daybar"><p>17</p></div>
+
<div class="daybar"><p>9</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
</ul>
</ul>
</div>
</div>
Line 1,971: Line 2,076:
<div class="open">
<div class="open">
<ul>
<ul>
 +
<li class="green e4"><p>Mini-prep from cultivated PompC+B0034+LacI+J61048 Eco.li&PSB1C3 E.coli</p></li>
 +
<li class="green e4"><p>digestion : [PompC+B0034+LacI+J61048]ES&[PSB1C3]EP</p></li>
 +
<li class="green e4"><p>Electrophoresis of [PompC+B0034+LacI+J61048] ES& [LacI+J61048]ES&[PSB1C3]EP.</p></li>
 +
<li class="green e4"><p>Single colony isolation from 37oCRBS + mGFP+Ter(B0015)</p></li>
</ul>
</ul>
</div>
</div>
Line 1,976: Line 2,085:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>18</p></div>
+
<div class="daybar"><p>10</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
</ul>
</ul>
</div>
</div>
Line 1,984: Line 2,096:
<div class="open">
<div class="open">
<ul>
<ul>
 +
<li class="green e3"><p>Mini-prep of cultivated 37oCRBS+mGFP+Ter(B0015)  E.coli</p></li>
 +
<li class="green e3"><p>Digestion:[ 37oCRBS+mGFP+Ter(B0015)]XP</p></li>
 +
<li class="green e3"><p>Single colony isolation from 37oCRBS+mGFP+Ter(B0015) LB-A plate, and cultivation in liquid LB-A.</p></li>
</ul>
</ul>
</div>
</div>
Line 1,989: Line 2,104:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>19</p></div>
+
<div class="daybar"><p>11</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
Line 1,999: Line 2,114:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>PCR of insert fragment [pLac+B0034+Zif268+AlsS] with designed primer for point mutation OK</p></li>
+
<li class="green e2"><p>PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)]</p></li>
-
<li class="green e2"><p>digestion: pLac+B0034+Zif268+AlsS [DPn1]</p></li>
+
<li class="green e2"><p>Transformation of mRFP on LB-K plate.</p></li>
</ul>
</ul>
</div>
</div>
Line 2,006: Line 2,121:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>20</p></div>
+
<div class="daybar"><p>12</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
Line 2,015: Line 2,130:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>transformation of pLac+B0034+Zif268+AlsS (point mutation), and cultivation on LB-A plate</p></li>
+
<li class="green"><p>Single colony isolation from mRFP LB-K plate, and cultivation in liquid LB-K</p></li>
</ul>
</ul>
</div>
</div>
Line 2,021: Line 2,136:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>21</p></div>
+
<div class="daybar"><p>13</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 2,030: Line 2,149:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>Single colony isolation from pLac+B0034+Zif268+AlsS (pm) LB-A plate, and cultivation in liquid LB-A</p></li>
+
<li class="green e5"><p>Ligation: insert[PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1C3]EP and insert[TetR]ES+[B0015]XP/vector[PSB1C3]EP.</p></li>
 +
<li class="green e5"><p>Mini-prep of cultivated mRFP E.coli</p></li>
 +
<li class="green e5"><p>Digestion:[mRFP]ES & [Ter(J61048)]XP & [37OCRBS+mGFP]XP</p></li>
 +
<li class="green e5"><p>Ligation: insert 37oCRBS+LuxR[ES]&37oCRBS+mGFP[XP]  /vector pSB1A3[EP]</p></li>
 +
<li class="green e5"><p>Transformation of 37oCRBS +LuxR+37oCRBS+mGFP on LB-A plate & mRFP +Ter(J61048) on LB-C plate& Ter(J61048) on LB-A plate.</p></li>
</ul>
</ul>
</div>
</div>
Line 2,036: Line 2,159:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>22</p></div>
+
<div class="daybar"><p>14</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
Line 2,046: Line 2,172:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>mini-prep of cultivated pLac+B0034+Zif268+AlsS (pm) E. coli </p></li>
+
<li class="green e5"><p>Transformation of PompC+B0034+LacI+J61048+PSB1C3 and TetR+B0015+PSB1C3</p></li>
-
<li class="green e2"><p>digestion : pLac+B0034+Zif268+AlsS (pm) [EP] </p></li>
+
<li class="green e5"><p>Cultivation of B0032+ho1 with liquid LB-A. PCR of insert fragment[Ter(B0015)]</p></li>
 +
<li class="green e5"><p>Single colony isolation from Ter(J61048) LB-A plate, and cultivation in liquid LB-A & 37OCRBS+mGFP on LB-K plate, and cultivation in liquid LB-K</p></li>
 +
<li class="green e5"><p>Transformation of 37oCRBS +LuxR+37oCRBS+mGFP on LB-A plate & mRFP +Ter(J61048) on LB-C plate</p></li>
 +
<li class="green e5"><p>Mini-prep of cultivated Ter (J61048)& 37oCRBS+mGFP E.coli</p></li>
</ul>
</ul>
</div>
</div>
Line 2,053: Line 2,182:
</div>
</div>
<div class="day brn">
<div class="day brn">
-
<div class="daybar"><p>23</p></div>
+
<div class="daybar"><p>15</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 2,062: Line 2,194:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>DNA sequencing OK</p></li>
+
<li class="green e4"><p>Digestion:[B0032]XP</p></li>
 +
<li class="green e4"><p>Transformation of  PompC+B0034+LacI+J61048 +PSB1C3and TetR+B0015 +PSB1C3 again</p></li>
 +
<li class="green e4"><p>Ligation: insert[PompC]ES+[B0032]XP/vector[PSB1K3]EP, then transformation of this part</p></li>
 +
<li class="green e4"><p>Mini-prep of B0032+ho1. PCR of insert fragment[37oCRBS + LuxR + 37oCRBS+mGFP]&[mRFP+Ter(J61048)].</p></li>
</ul>
</ul>
</div>
</div>
Line 2,070: Line 2,205:
<!---------------------------------------- week end ---------------------------------------->
<!---------------------------------------- week end ---------------------------------------->
<!---------------------------------------- week start ---------------------------------------->
<!---------------------------------------- week start ---------------------------------------->
-
<div class="week">
+
<div class="week bigwk">
<div class="day">
<div class="day">
-
<div class="daybar"><p>24</p></div>
+
<div class="daybar"><p>16</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
</ul>
</ul>
</div>
</div>
Line 2,080: Line 2,216:
<div class="open">
<div class="open">
<ul>
<ul>
 +
<li class="green"><p>PCR + single colony isolation of PompC+B003+LacI+J61048 Cr E.coli&TetR+B0015 Cr E.coli&PompC+B0032 Kr E.coli, then Electrophoresis of  these parts</p></li>
</ul>
</ul>
</div>
</div>
Line 2,085: Line 2,222:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>25</p></div>
+
<div class="daybar"><p>17</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
<li class="caldot green"></li>
<li class="caldot green"></li>
 +
</ul>
 +
</div>
 +
 +
<div class="open">
 +
<ul>
 +
<li class="green"><p>Mini-prep of cultivated PompC+B0034 E.coli and TetR+B0015 Cr E.coli </p></li>
 +
</ul>
 +
</div>
 +
 +
</div>
 +
<div class="day">
 +
<div class="daybar"><p>18</p></div>
 +
<div class="dots">
 +
<ul>
 +
</ul>
 +
</div>
 +
 +
<div class="open">
 +
<ul>
 +
</ul>
 +
</div>
 +
 +
</div>
 +
<div class="day">
 +
<div class="daybar"><p>19</p></div>
 +
<div class="dots">
 +
<ul>
 +
</ul>
 +
</div>
 +
 +
<div class="open">
 +
<ul>
 +
</ul>
 +
</div>
 +
 +
</div>
 +
<div class="day">
 +
<div class="daybar"><p>20</p></div>
 +
<div class="dots">
 +
<ul>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 2,095: Line 2,272:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>PCR of insert fragment [B0034+ PBSII+ilvC] with designed primer for point mutation OK</p></li>
+
<li class="green"><p>Ligation: Pcons+RBS+pcyA&B0032+ho1&pSB1A3.</p></li>
-
<li class="green e2"><p>digestion : B0034+ PBSII+ilvC [DPn1]</p></li>
+
</ul>
 +
</div>
 +
 
 +
</div>
 +
<div class="day">
 +
<div class="daybar"><p>21</p></div>
 +
<div class="dots">
 +
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
</ul>
 +
</div>
 +
 
 +
<div class="open">
 +
<ul>
 +
<li class="green e6"><p>Ligation: insert[PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1A3]EP</p></li>
 +
<li class="green e6"><p>Transformation of Pcons+RBS+pcyA+B0032+ho1 on LB-A plate. PCR of insert fragment[37oCRBS + LuxR + 37oCRBS+mGFP]</p></li>
 +
<li class="green e6"><p>Digestion:[pSB1C3]EP&[mRFP]ES&[Ter(J61048)]XP</p></li>
 +
<li class="green e6"><p>Ligation: insert mRFP[ES]&Ter(J61048)[XP]  /vector pSB1C3[EP]</p></li>
 +
<li class="green e6"><p>Transformation of mRFP + Ter (J61048) on LB-C plate</p></li>
 +
<li class="green e6"><p>Single colony isolation from 37oCRBS + LuxR +37oCRBS+mGFP LB-A plate, and cultivation in liquid LB-A.</p></li>
</ul>
</ul>
</div>
</div>
</div>
</div>
 +
<div class="day brn">
 +
<div class="daybar"><p>22</p></div>
 +
<div class="dots">
 +
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
</ul>
 +
</div>
 +
 +
<div class="open">
 +
<ul>
 +
<li class="green e3"><p>Transformation of PompC+B0034+LacI+J61048+PSB1A3 but there is no colony appearing. PCR of insert fragment[mRFP+Ter(J61048)]</p></li>
 +
<li class="green e3"><p>Mini-prep of cultivated  37o CRBS + LuxR + 37oCRBS+mGFP E.coli</p></li>
 +
<li class="green e3"><p>Digestion:[ 37o CRBS + LuxR + 37oCRBS+mGFP]ES</p></li>
 +
</ul>
 +
</div>
 +
 +
</div>
 +
</div>
 +
<!---------------------------------------- week end ---------------------------------------->
 +
<!---------------------------------------- week start ---------------------------------------->
 +
<div class="week bigwk">
<div class="day">
<div class="day">
-
<div class="daybar"><p>26</p></div>
+
<div class="daybar"><p>23</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 2,111: Line 2,340:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>transformation of B0034+ PBSII+ilvC (pm), and cultivation on LB-C plate------Fail</p></li>
+
<li class="green e6"><p>Digestion:[PompC+B0032]ES and [TetR+B0015]XP,[LacI+J61048]XP</p></li>
 +
<li class="green e6"><p>Ligation: insert [PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1C3]EP</p></li>
 +
<li class="green e6"><p>Transformation  of PompC+B0034+LacI+J61048 +PSB1C3</p></li>
 +
<li class="green e6"><p>Electrophoresis of [PompC+B0032]ES & [TetR+B0015]XP</p></li>
 +
<li class="green e6"><p>Digestion again:[PompC+B0032]ES&[LacI+J61048]XP&[LacI+B0015]XP</p></li>
 +
<li class="green e6"><p>Transformation of mRFP + Ter (J61048) on LB-C plate</p></li>
 +
</ul>
 +
</div>
 +
 
 +
</div>
 +
<div class="day">
 +
<div class="daybar"><p>24</p></div>
 +
<div class="dots">
 +
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
</ul>
 +
</div>
 +
 
 +
<div class="open">
 +
<ul>
 +
<li class="green e7"><p>PCR and single colony isolation of  PompC+B0034+LacI+J61048+PSB1C3 E.coli</p></li>
 +
<li class="green e7"><p>Electrophoresis of [B0032]dig ES&[TetR+B0015]dig XP&[LacI+J61048]dig XP&PCR product [PompC+B0034+LacI+J61048] ,[TetR+B0015] OK</p></li>
 +
<li class="green e7"><p>Decide to determine the sequence of [TetR+B0015] OK.</p></li>
 +
<li class="green e7"><p>Single colony isolation from 6/21 plate.</p></li>
 +
<li class="green e7"><p>Cultivation of Pcons+ RBS+pcyA+B0032+ho1 with liquid LB-A.</p></li>
 +
<li class="green e7"><p>Electrophoresis of Pcons+RBS+PcyA+B0032+ho1.</p></li>
 +
<li class="green e7"><p>PCR of insert fragment[mRFP+Ter(J61048)].</p></li>
 +
</ul>
 +
</div>
 +
 
 +
</div>
 +
<div class="day">
 +
<div class="daybar"><p>25</p></div>
 +
<div class="dots">
 +
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
</ul>
 +
</div>
 +
 
 +
<div class="open">
 +
<ul>
 +
<li class="green e7"><p>Ligation:insert[PompC+B0034]ES+[LacI+J61048]XP&[PompC]ES+[B0032]XP/vector[PSB1C3]EP and [PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1A3]EP</p></li>
 +
<li class="green e7"><p>Transformation of the ligation product made today</p></li>
 +
<li class="green e7"><p>PCR of Pcons+RBS+pcyA+B0032+ho1.</p></li>
 +
<li class="green e7"><p>Cultivation of Pcons+RBS+pcyA+B0032+ho1 with liquid LB-A</p></li>
 +
<li class="green e7"><p>Electrophoresis of Pcons+RBS+pcyA+B0032+ho1.</p></li>
 +
<li class="green e7"><p>Mini-prep of Pcons+RBS+pcyA+B0032+ho1.</p></li>
 +
<li class="green e7"><p>Ligation: insert mRFP[ES]&Ter(J61048)[XP]/vector pSB1C3[EP]</p></li>
</ul>
</ul>
</div>
</div>
Line 2,117: Line 2,405:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>27</p></div>
+
<div class="daybar"><p>26</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 2,126: Line 2,418:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>PCR of insert fragment [B0034+ PBSII+ilvC] with designed primer for point mutation OK</p></li>
+
<li class="green e5"><p>PCR and single colony isolation of PompC+B0034+LacI+J61048+PSB1A3& PompC+B0034+LacI+J61048+PSB1C3&PompC+B0032+PSB1C3 E.coli</p></li>
 +
<li class="green e5"><p>Electrophoresis of the PCR product made today</p></li>
 +
<li class="green e5"><p>Electrophoresis of Pcons+RBS+pcyA+B0032+ho1──NOT OK</p></li>
 +
<li class="green e5"><p>Ligation: Pcons+RBS+pcyA&B0032+ho1</p></li>
 +
<li class="green e5"><p>Transformation of Pcons+RBS+pcyA+B0032+ho1. Transformation of mRFP + Ter (J61048) on LB-C plate</p></li>
</ul>
</ul>
</div>
</div>
Line 2,132: Line 2,428:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>28</p></div>
+
<div class="daybar"><p>27</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 2,141: Line 2,441:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>digestion : B0034+ PBSII+ilvC [DPn1]</p></li>
+
<li class="green e5"><p>Cultivation of PompC+B0034+LacI+J61048+PSB1A3 E.coli in liquid LB-A tubes</p></li>
 +
<li class="green e5"><p>PCR of Pcons+RBS+pcyA+B0032+ho1</p></li>
 +
<li class="green e5"><p>Cultivation of Pcons+RBS+pcyA+B0032+ho1 with liquid LB-A</p></li>
 +
<li class="green e5"><p>Electrophoresis of Pcons+RBS+pcyA+B0032+ho1──NOT OK</p></li>
 +
<li class="green e5"><p>Transformation of mRFP + Ter (J61048) on LB-C plate</p></li>
</ul>
</ul>
</div>
</div>
Line 2,147: Line 2,451:
</div>
</div>
<div class="day">
<div class="day">
-
<div class="daybar"><p>29</p></div>
+
<div class="daybar"><p>28</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
</ul>
</ul>
Line 2,156: Line 2,462:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green"><p>transformation of B0034+ PBSII+ilvC (pm), and cultivation on LB-C plate</p></li>
+
<li class="green e3"><p>PCR of Pcons+RBS+pcyA+B0032+ho1</p></li>
 +
<li class="green e3"><p>Electrophoresis of Pcons+RBS+pcAa+B0032+ho1──NOT OK</p></li>
 +
<li class="green e3"><p>Ligation: insert mRFP[ES]&Ter(J61048)[XP]  /vector pSB1C3[EP]</p></li>
</ul>
</ul>
</div>
</div>
Line 2,162: Line 2,470:
</div>
</div>
<div class="day brn">
<div class="day brn">
 +
<div class="daybar"><p>29</p></div>
 +
<div class="dots">
 +
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
</ul>
 +
</div>
 +
 +
<div class="open">
 +
<ul>
 +
<li class="green e8"><p>Cultivation of PompC+B0034+LacI+J61048+PSB1A3 E.coli in liquid LB-A tubes again. </p></li>
 +
<li class="green e8"><p>Ligation: Pcons+RBS+pcyA&B0032+ho1&pSB1A3</p></li>
 +
<li class="green e8"><p>Transformation of ligated Pcons+RBS+pcyA+B0032+ho1</p></li>
 +
<li class="green e8"><p>Single colony isolation from 6/21 Pcons+RBS+pcyA+B0032+ho1 LB plate</p></li>
 +
<li class="green e8"><p>PCR of 6/21 Pcons+RBS+pcyA+B0032+ho1</p></li>
 +
<li class="green e8"><p>Electrophoresis of 621 Pcons+RBS+pcyA+B0032+ho1─NOT OK</p></li>
 +
<li class="green e8"><p>Transformation of mRFP+Ter(J61048) on LB-C plate</p></li>
 +
<li class="green e8"><p>Single colony isolation from 37oCRBS +LuxR +37oCRBS+mGFP LB-A plate, and cultivation in liquid LB-A</p></li>
 +
</ul>
 +
</div>
 +
 +
</div>
 +
</div>
 +
<!---------------------------------------- week end ---------------------------------------->
 +
<!---------------------------------------- week start ---------------------------------------->
 +
<div class="week bigwk">
 +
<div class="day">
<div class="daybar"><p>30</p></div>
<div class="daybar"><p>30</p></div>
<div class="dots">
<div class="dots">
<ul>
<ul>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
 +
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
<li class="caldot green"></li>
Line 2,172: Line 2,519:
<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>PCR of insert fragment [B0034+ PBSII+ilvC] (pm) OK</p></li>
+
<li class="green e8"><p>Mini-prep of cultivated PompC+B0034+LacI+J61048+PSB1A3 E.coli and then digest with EcoR1 and SpeI:[ PompC+B0034+LacI+J61048]dig ES</p></li>
-
<li class="green e2"><p>Single colony isolation from B0034+ PBSII+ilvC (pm) LB-C plate, and cultivation in liquid LB-C</p></li>
+
<li class="green e8"><p>Electrophoresis of [PompC+B0034+LacI+J61048+PSB1A3] mini and [ PompC+B0034+LacI+J61048]dig ES OK</p></li>
 +
<li class="green e8"><p>Decide to determine 
 +
sequence of[ PompC+B0034+LacI+J61048]</p></li>
 +
<li class="green e8"><p>Cultivation of 6/29 Pcons+RBS+PcyA+B0032+ho1 LB plate with liquid LB-A</p></li>
 +
<li class="green e8"><p>PCR of 6/29 Pcons+RBS+PcyA+B0032+ho1</p></li>
 +
<li class="green e8"><p>Cultivation of 6/29 re-ligated Pcons+RBS+PcyA+B0032+ho1</p></li>
 +
<li class="green e8"><p>PCR of 6/29 re-ligated Pcons+RBS+pcyA+B0032+ho1</p></li>
 +
<li class="green e8"><p>Electrophoresis of all above─NOT OK</p></li>
 +
</ul>
 +
</div>
 +
 
 +
</div>
 +
<div class="day">
 +
<div class="daybar"><p></p></div>
 +
<div class="dots">
 +
<ul>
 +
</ul>
 +
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 +
 
 +
<div class="open">
 +
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 +
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 +
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 +
 
 +
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 +
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 +
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 +
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 +
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 +
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 +
 
 +
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 +
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 +
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 +
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 +
 
 +
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 +
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 +
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 +
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 +
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 +
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 +
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 +
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 +
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 +
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 +
 
 +
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<div class="open">
<div class="open">
<ul>
<ul>
-
<li class="green e2"><p>mini-prep of cultivated B0034+ PBSII+ilvC (pm) E. coli </p></li>
+
<li class="green"><p>PCR + single colony isolation of PompC+B003+LacI+J61048 Cr E.coli&TetR+B0015 Cr E.coli&PompC+B0032 Kr E.coli, then Electrophoresis of  these parts</p></li>
-
<li class="green e2"><p>digestion : B0034+ PBSII+ilvC (pm) [EP] </p></li>
+
</ul>
</ul>
</div>
</div>

Revision as of 09:12, 21 September 2013

Notes

The experimental log that records down the purpose, method, result and date of each experiment that we have conducted.

About the notes

Our notes are the abbreviations of the experimental process, including titles, genes we transformed, plasmids we used, and resistances on the plasmids. We categorized these notes by date. Moreover, team members followed the iGEM protocol in each experimental step in order to get better experimental efficiency and the results we had expected.

March 2013

Sunday

Monday

Tuesday

Wednesday

Thursday

Friday

Saturday

24

25

26

27

28

29

  • Transformation of ho1(Kr), Cph8(Cr), 32℃ RBS(Ar) and cultivation on three LB plates.

  • transformation of PompC and cultivation on LB-A plate

30

  • Single colony isolation from three plates and cultivation them in liquid LB

  • Single colony isolation from PompC LB-C plate and cultivation in liquid LB-A

  • mini-prep of cultivated PompC E.coli

31

  • Transformation of RBS B0030+PSB1A3&B0034+PSB1A3 and cultivation on LB-A plate.

  • Mini-prep for cultivated ho1(Kr), Cph8(Cr), 32℃ RBS(Ar) PCR of ho1(Kr), Cph8(Cr), 32℃ RBS(Ar).

  • Electrophoresis of ho1(Kr), Cph8(Cr), 32℃ RBS(Ar) ──NOT OK.

  • PCR of insert fragment [PompC+psB1A2]

April 2013

Sunday

Monday

Tuesday

Wednesday

Thursday

Friday

Saturday

1

  • Cultivation of B0030&B0034 Ar E.coli in liquid LB-A tubes.

2

  • Mini-prep of cultivated B0030&B0034 E.coli and then digestion: [B0030]XP&[B0034]XP.

3

4

5

6

  • Electrophoresis of [B0030]XP&[B0034]XP Not OK.

  • PCR of ho1 to check ho1 and transform to test the resistance.

7

  • Electrophoresis of [B0030]mini&dig XP and [B0034]mini&dig XP OK

  • Cultivation of B0030&B0034 Ar E.coli in liquid LB-A tubes.

  • Cultivation of ho1 in liquid LB-K and LB-K plate

  • Cultivation of Cph8+RBS in liquid LB-C.

    • 8

    • Mini-prep of cultivated B0030&B0034 Ar E.coli. Mini-prep of ho1 & Cph8+RBS.

    9

    • Electrophoresis of mini ho1&Cph8+RBS.

    10

    11

    • PCR of mini ho1&Cph8+RBS

    • Electrophoresis of ho1&Cph8+RBS(After PCR)

    • Digestion: [ho1]EP&[Cph8+RBS]EP.

    12

    • Transformation of Plac&Ptet& pcyA but there is no colony appearing in pcyA plate.

    • Electrophoresis of digested ho1&Cph8+RBS (NOT OK).

    13

    14

    • Cultivation of Ptet&Plac E.coli in LB tubes.

    • Transformation: LuxR,B0015,J61048 and cultivation on LB-A plate

    15

    • Mini-prep of cultivated Ptet&Plac E.coli

    • Digestion:[Ptet]ES&[Plac]ES PCR of insert fragment pcyA

    • Electrophoresis of [Ptet]dig ES&[pcyA]PCR&[Plac]dig ES.

    • Single colony isolation from J61048,B0015,LuxR plate and cultivation in liquid LB-A mini-prep of cultivated LuxR,J61048,B0015

    16

    • PCR of mini Cph8+RBS

    • Electrophoresis of Cph8+RBS(After PCR)──NOT OK.

    • digestion: [J61048]EP,[B0015]EP,[LuxR]EP

    • Electrophoresis of ter. mini [J61048] dig E/EP and t er. mini [B0015] dig E/EP and luxR mini dig E/EP

    17

    • PCR of mini Cph8+RBS Electrophoresis of Cph8+RBS(After PCR)──NOT OK.

    • Electrophoresis of the products of digestion of ter. [J61048], ter. [B0015] , luxR

    • transformation of tetR plasmid and cultivation on LB –A plate 3. Digestion: ter. [J61048], ter. [B0015] dig EP

    18

    19

    • Cultivation of pcyA Kr E.coli on LB-K plates

    • mini-prep of cultivated tetR E.coli

    • digestion: tetR+pSB1A2 dig EP

    • electrophoresis of tetR dig EP,ter.[J61048 ] dig EP and ter.[B0015] dig EP

    20

    • Cultivation of pcyA Kr E.coli in liquid LB-K tubes.

    • PCR of tetR,ter.[J61048] and ter.[B0015 ]mini

    • electrophoresis of PCR products of tetR mini,J61048 mini and B0015 mini

    21

    • Single colony isolation of pcyA Kr E.coli

    • Mini-prep of cultivated pcyA Kr E.coli

    • Digestion:[pcyA]ES

    • Electrophoresis of [pcyA]mini&dig ES OK

    • PCR of tetR,ter.[J61048] and ter.[B0015 ]mini

    • electrophoresis of PCR products of tetR mini,J61048 mini and B0015 mini

    • electrophoresis of the digestion products of tetR dig EP and B0015 dig EP

    • aliquot every 20ul of PompC,J61048 and LuxR

    22

    23

    24

    25

    26

    27

    28

    • Transformation of PompC mini and cultivation on LB-A plate

    29

    • transformation of PompC , Ar mini and cultivation on LB-A plate

    • single colony isolation from PompC , Ar

    30

    • Transformation of 37ﾟC RBS and ho1.

    • Mini-prep of cultivated PompC , Ar E.coli

    May 2013

    Sunday

    Monday

    Tuesday

    Wednesday

    Thursday

    Friday

    Saturday

    1

    • Cultivation of 37ﾟC RBS and ho1

    • Digestion: [pSB1C3] EP&[B0030]ES&{pcyA]XP

    • Transformation of pSB1A3 &pSB1K3

    • Transformation of mGFP[pSB1A2] and cultivation on LB-A plate.

    2

    • Cultivation of PompC&B0034&LacI&Plac&B0030&TetR&J61048&B0015 Ar E.coli in liquid LB-A tubes.

    • Cultivation of pSB1A3&pSB1K3 with liquid LB

    • Mini-prep of 37ﾟC RBS and ho1──Fail

    • Ligation: [pSB1C3] EP&[B0030]ES& {pcyA]XP

    • Transformation of B0030

    • Cultivation of 37ﾟC RBS and ho1 with liquid LB

    • Transformation of RBS+pcyA+psB1C3.

    • Transformation of mGFP[pSB1A2] and cultivation on LB-A plate

    3

    • Mini-prep of cultivated PompC &B0034&LacI&Plac&B0030&TetR&J61048&B0015,and then digestion:[PompC ]ES&[B0034]XP&[LacI]ES&[Plac]XP&[B0030]ES&[TetR]ES&[TetR]XP&[J61048]XP&[J61048]ES&[B0015]XP.

    • Mini-prep of ho1,pSB1A3, pSB1K3 and 37ﾟC RBS

    • Digestion: [B0030+pcyA]XP

    • Cultivation of B0030 with liquid LB-C

    • Check RBS+pcyA+psB1C3(PCR & Electrophoresis)──FAIL

    • Digestion: [Pcons]ES& [ho1]XP.

    • Single colony isolation from 37。C RBS Ar,pSB1A3,pSB1C3 and pSB1K3

    • electrophoresis of mGFP dig E

    4

    • Electrophoresis of [PompC]ES&[B0030]ES&[LacI]ES&[TetR]ES&[J61048]XP&[TetR]ES&[B0015]XP OK

    • Ligation: insert [PompC]ES+[B0030]XP&[LacI]ES+[J61048]XP&[J61048]ES+[Plac]XP&[B0030]ES+[TetR]XP&[TetR]ES+[B0015]XP/vector[PSB1C3]EP

    • Transformation of B0030+ho1.

    • Mini-prep of cultivated 37。C RBS Ar pSB1A3,37。C RBS+mGFP, Kr and pSB1K3 E.coli

    • digestion:37。C RBS dig ES,mGFP dig XP,LuxR dig XP and pSB1K3 dig EP

    • electrophoresis of digestion products of LuxR,37。C RBS, mGFP and pSB1K3

    • ligation:37。C RBS+luxR+pSB1K3

    • transformation of 37。C RBS+mGFP, Kr->pSB1K3 and cultivation on LB-K plate

    5

    • Electrophoresis of digested RBS+pcyA──OK

    • Ligation: RBS+ho1

    • Cultivation of J61048 with liquid LB-C

    • Digestion: [pSB1A3]EP &[pSB1K3]EP

    • Transformation of ligated RBS+ho1.

    • Ligation:37。C RBS+luxR+pSB1K3 and 37。C RBS+mGFP+pSB1K3

    • transformation of 37。C RBS+luxR, Kr and 37。C RBS+mGFP ,Kr and cultivation on LB-K plate

    6

    • Mini-prep of J61048

    • Ligation: RBS+pcyA+Pcons&pSB1K3

    • Electrophoresis of J61048.

    • Digestion: [J61048]XP

    • Transformation of RBS+pcyA+Pcons.

    • Check LB-A&C&K plates.

    7

    • Mini-prep of J61048 -Ligation: RBS+pcyA+Pcons&pSB1K3

    • Electrophoresis of J61048.

    • Digestion: [J61048]XP

    • Transformation of RBS+pcyA+Pcons.-Check LB-A&C&K plates.

    8

    • Ligation:insert[PompC]ES+[B0034]XP&[LacI]ES+[J61048]XP&[B0030]ES+[TetR]XP&[TetR]ES+[B0015]XP/vector[PSB1C3]E

    • PCR of 37。RBS+luxR, Kr and 37。C RBS+mGFP ,Kr

    • electrophoresis of 37。RBS+luxR ,Kr and 37。C RBS+mGFP ,Kr

    9

    • Transformation and cultivation of PompC+B0034&LacI+J61048&B0030+TetR&TetR+B0015(PSB1C3) on LB-C plates.

    • PCR of 37。RBS+luxR, Kr and 37。C RBS+mGFP ,Kr

    • electrophoresis of 37。RBS+luxR, Kr and 37。C RBS+mGFP ,Kr

    • single colony isolation from37。RBS+luxR, Kr and 37。C RBS+mGFP ,Kr

    10

    • Cultivation of Pcons+RBS+pcyA and B0030 with liquid LB.

    • Mini-prep of cultivated 37。C RBS+luxR, Kr E.coli and 37。C RBS+mGFP ,Kr E.coli

    • digestion: 37。RBS+luxR, dig ES and 37。C RBS+mGFP dig ES

    11

    • Mini-prep of Pcons+RBS+pcyA.

    • Electrophoresis of Pcons+RBS+pcyA──NOT OK.

    • Electrophoresis of 37。C RBS+mGFP, Kr mini, 37。C RBS+mGFP dig ES, 37。C RBS+luxR, Kr mini and 37。C RBS+luxR dig ES

    12

    • Electrophoresis of 37。C+luxR mini, dig ES and 37。C RBS+mGFP mini,dig ES

    13

    • Cultivation of TetR+B0015 Cr E.coli in liquid LB-C tube

    • Single colony isolation of backbone PSB1K3 E.coli

    • Ligation:insert[B0030]ES+[TetR]XP&[TetR]ES+[B0015]XP/vector[PSB1C3]EP.

    • Electrophoresis of Pcons+RBS+pcyA──NOT OK

    • Transformation of B0032&B0034.

    14

    • Ligation:insert[PompC]ES+[B0034]XP&[LacI]ES+[J61048]XP/vector[PSB1C3]EP.

    • Cultivation of Pcons+RBS+pcyA & B0032&B0034 with liquid LB.

    15

    • Mini-prep of cultivated PSB1K3 E.coli

    • Cultivation of B0015+TetR+PSB1C3 and TetR+B0015+PSB1C3 in liquid LB-C plates and Plac+PSB1A3 in liquid LB-A plate. Mini-prep of Pcons+RBS+pcyA&B0032&B0034.

    16

    • Ligation:insert[PompC]ES+[B0034]XP&[LacI]ES+[J61048]XP/vector[PSB1K3]EP

    • Transformation and cultivation of PompC+B0034&LacI+J61048 (PSB1K3) on LB-K plates.

    17

    • cultivation of PompC+B0034&LacI+J61048 Kr E.coli on liquid LB-K tubes.

    • single colony isolation of PompC+B0034&LacI+J61048 Kr E.coli.

    • Electrophoresis of Pcons+RBS+pcyA(NOT OK) &B0032&B0034

    • Cultivation of Pcons+RBS+pcyA.Digestion: [B0032]ES &[B0023]ES.

    18

    • Cultivation of PompC+B0034&LacI+J61048 Kr E.coli from single colony isolation plate made yesterday

    • Min-prep of cultivated [B0030+TetR]Cr&[ Plac]Ar&[PompC+B0034]Kr&[LacI+J61048]Kr E.coli and then digestion:[PompC+B0034]ES &[LacI+J61048]XP. Mini-prep Pcons+RBS+pcyA

    • Electrophoresis of Pcons+RBS+pcyA& digested B0032&B0034──NOT OK

    • Digestion: [B0032]ES &[B0034]ES &[Pcons+RBS+pcyA]E.

    19

    • Electrophoresis of [LacI+J61048]mini & dig XP and [PompC+B0034]mini & dig ES and [PSB1K3]mini & dig EP.

    • Electrophoresis of digested B0032&B0034&Pcons+RBS+pcyA──OK

    • Transformation of pSB1C3

    20

    • Digestion:[Plac]P.

    • Ligation: Pcons+RBS+pcyA&pSB1K3, B0032+ho1&pSB1C3, B0034+ho1&pSB1C1

    • Transformation of Pcons+RBS+pcyA&pSB1K3, B0032+ho1&pSB1C3, B0034+ho1&pSB1C1

    • Single colony isolation from pSB1C3

    21

    • Single colony isolation of TetR+B0015 Cr Ecoli

    • Electrophoresis of [PompC+B0034]mini&dig ES and[Plac]mini&dig P.

    • Cultivation of Pcons+RBS+pcyA &B0032+ho1 with liquid LB

    • Mini-prep of cultivated pSB1C3 E.coli

    22

    • Digestion:[Plac]XP

    • Electrophoresis of [PompC+B0034]mini&dig ES and[Plac]mini&dig XP

    • Ligation: insert[TetR]ES+[B]XP/vector[PSBC3]EP

    • Mini-prep of Pcons+RBS+pcyA

    • Mini-prep of cultivated 37。RBS+mGFP E.coli

    • digestion:37。C RBS+mGFP dig ES and mini

    • electrophoresis of 37。C RBS+mGFP mini and 37。C RBS+mGFP dig ES.

    23

    • Single colony isolation of TetR+B0015 transformation of TetR +B0015 +PSB1C3 on LB-C plates

    • Ligation +transformation on: insert[TetR]ES+[B0015]XP&[PompC]ES+[B0034]/vector[PSB1K3]EP on LB-K plates

    24

    • Single colony isolation of PompC+B0034+PSB1K3 E.coli

    • Electrophoresis of Pcons+RBS+pcyA.

    • -Ligation: B0032+ho1&B0034+ho1.

    • Digestion; [Pcons+RBS+pcyA]ES &[ho1]XP&[pSB1C3]EP.

    • Electrophoresis of 37。 RBS+mGFP dig ES

    25

    • Ligation and transformation: insert [PompC]ES+[B0034]XP/vector[PSB1K3]EP on LB-K plates

    • Single colony isolation of PompC+B0034+PSB1K3 E.coli again

    • Mini-prep of cultivated TetR+B0015 Cr E.coli and digestion:[TetR+B0015]P

    • Electrophoresis of :[TetR+B0015]P

    • Digestion: [TetR+B0015]XP

    • Cultivation of PompC+B0034 in liquid LB-K tubes.

    • Mini-prep of B0032+ho1

    • Cultivation of B0034+ho1 with liquid LB-A

    26

    • Cultivation of PompC+B0034Kr E.coli in liquid LB-K tubes again

    • Mini-prep of cultivate PompC+B0034 Kr E.coli

    • Electrophoresis of [TetR+B0015] dig XP not OK

    • Mini-prep of B0034+ho1 Electrophoresis of Pcons+RBS+pcyA & B0032+ho1 & B0034+ho1.

    27

    • Digestion:[TetR+B0015]XP

    • Electrophoresis of [TetR+B0015]mini&dig XP

    • Digestion:[PompC+B0034]E first

    • Electrophoresis of [PompC+B0034]E

    • Digestion: [PompC+B0034]E, S later

    • Ligation: insert[PompC]ES+[B0034]XP/vector[PSB1K3]EP

    28

    • Electrophoresis of [PompC+B0034]ES not OK

    • Transformation of PompC+B0034+PSB1K3.

    • Electrophoresis of B0034+ho1

    • Digestion: [B0034+ho1]XP &[Pcons+RBS+pcyA]E

    29

    • PCR and electrophoresis test. Electrophoresis of digested B0034+ho1 &Pcons+RBS+pcyA.

    30

    • Cultivation of PompC+B0034+PSB1K3 E.coli in liquid LB-K tubes

    • Ligation: insert[TetR]ES+[B0015]XP/vector[PSB1K3]EP

    • Transformation of Plac+PSB1A3 on LB-A plate

    • Digestion:B0015 Ar dig XP, pSB1A3 Ar dig EP and pSB1C3 Cr dig EP

    • ligation:37。C RBS+mGFP+ter. [B0015], Cr

    • transformation of 37。C RBS+mGFP+ter., Cr

    31

    • Mini-prep of cultivated PompC+B0034 Kr E.coli, then

    • Digestion and electrophoresis of [PompC+B0034]E

    • Decide to determine the sequence of PompC+B0015.

    • Cultivation of ho1 with liquid LB-K and pSB1C3 with liquid LB-C.

    • PCR of 37。C RBS+mGFP+ter. Cr

    • electrophoresis of 37。C RBS+mGFP+ter., ter.[B0015] digXP, pSB1C3 dig EP and pSB1A3 dig EP

    • transformation of BBa _E1010 Kr

    June 2013

    Sunday

    Monday

    Tuesday

    Wednesday

    Thursday

    Friday

    Saturday

    1

    • PCR and single colony isolation of TetR+B0015+PSB1K3

    • Electrophoresis of PCR product made at this morning and Plac(5/30 Ar) OK

    • Digestion:[Ter(B0015)]XP

    • Transformation of Ter(B0015) on LB-A plate & mGFP on LB-K plate & 37oC RBS+mGFP+Ter(B0015) on LB-C plate

    • Ligation: insert 37oC RBS[ES] & Ter(B0015)[XP]/ vector pSB1C3[EP]

    • Single colony isolation from Ter(B0015) LB-A plate, and cultivation in liquid LB-A

    2

    • Ligation: insert [TetR]ES+[B0015]XP/vector [PSB1K3]EP and transformation of this part.

    • Ligation: B0032+ho1&B0034+ho1

    • Transformation of B0032+ho1&B0034+ho1

    • Cultivation of pSB1C3(C20&C10)with liquid LB.

    • PCR of insert fragment[37oC RBS+mGFP+Ter(B0015)]

    • Transformation of mRFP on LB-A plate& LB-C plate& LB-K plate-Mini-prep of cultivated Ter(B0015) E.coli.

    3

    • PCR of Plac &TetR+B0015

    • Mini-prep of pSB1C3

    • Cultivation of B0032+ho1&B0034+ho1 with LB-A plate PCR of B0032+ho1&B0034+ho1

    • Electrophoresis of B0032+ho1&B0034+ho1──NOT OK.

    • Single colony isolation from 37oC RBS+mGFP+Ter(B0015) LB-K plate, and cultivation in liquid LB-K & mRFP LB-A, and cultivation in liquid LB-K

    • Mini-prep of cultivated 37oC RBS+mGFP+Ter(B0015) E.coli

    • Digestion:[37oC RBS+mGFP+Ter(B0015)]XP

    4

    • Electrophoresis of Plac .TetR+B0015 OK

    • Cultivation of Plac.TetR+B0015 in liquid LB-A tubes. PCR of B0032+ho1&B0034+ho1

    • Electrophoresis of B0032+ho1(OK)&B0034+ho1(After PCR)

    • Mini-prep of B0032+ho1. Digestion: [B0032+ho1]XP.

    • Cultivation of ho1 with liquid LB-K

    • Single colony isolation from mRFP LB-A plate, and cultivation in liquid LB-A

    • Digestion:[37oC RBS+mGFP+Ter(B0015)]XP

    • PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)].

    5

    • Mini-prep of cultivated Plac and TetR+B0015 E.coli and then conduct digestion :[Plac]P&[TetR+B0015]P

    • Electrophoresis of [Plac ]P and [TetR+B0015]P,[Plac]XP and [TetR+B0015]XP

    • Ligation: insert[PompC+B0034]ES+[LacI+J61048]XP/vector [PSB1A3]EP

    • Electrophoresis of B0032+ho1 mini&B0032+ho1(digested)&pSB1C3 mini&pSB1C3(digested)&ho1 mini&ho1(digested)

    • Mini-prep of ho1

    • Digestion: [ho1]XP

    • Digestion:[37oC RBS+mGFP+Ter(B0015)]XP

    • Transformation of mRFP on LB-A plate

    6

    • Transformation of PompC+B0034+LacI+J61048+PSB1A3

    • Ligation: insert [PompC+B0034]ES+[LacI+J61048]XP/vector [PSB1A3]EP

    • Electrophoresis of B0032+ho1 mini&B0032+ho1(digested)&ho1 mini&ho1 (digested).

    • Single colony isolation from mRFP LB-A plate, and cultivation in liquid LB-A & LB-K & LB

    • Digestion:[Ter(B0015)]EX

    • Ligation: insert 37oC RBS+mGFP[ES]&Ter(B0015)[EX]

    • Transformation of 37oCRBS+mGFP+Ter(B0015) on LB-K plate & LB-A plate, mRFP on LB-A plate

    7

    • Transformation of PompC+B0034+LacI+J61048 again

    • Single colony isolation of TetR+B0015 Kr E.coli

    • Digestion:[LacI+J61048]ES. PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)]

    • Single colony isolation from 37oCRBS+mGFP+Ter(B0015) LB-A plate, and cultivation in liquid LB-A.

    8

    • PCR + single colony isolation of PompC+B0034+LacI+J61048 E.coli

    • Electrophoresis of PompC+B0034+LacI+J61048 OK

    • Ligation: insert [LacI+J61048]ES+[Plac]XP/vector[PSB1C3]EP

    • Transformation of LacI+J61048+Plac on LB-C plate

    • Cultivation of PompC+B0034+LacI+J61048 in LB-A tubes & TetR+B0015 in LB-K tube & PSB1A3 in LB-A tube &PSB1C3 in LB-C tube.

    • Mini-prep of cultivated 37oCRBS+mGFP+Ter(B0015) E.coli

    • Digestion:[ 37oCRBS+mGFP+Ter(B0015)]XP

    • PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)].

    9

    • Mini-prep from cultivated PompC+B0034+LacI+J61048 Eco.li&PSB1C3 E.coli

    • digestion : [PompC+B0034+LacI+J61048]ES&[PSB1C3]EP

    • Electrophoresis of [PompC+B0034+LacI+J61048] ES& [LacI+J61048]ES&[PSB1C3]EP.

    • Single colony isolation from 37oCRBS + mGFP+Ter(B0015)

    10

    • Mini-prep of cultivated 37oCRBS+mGFP+Ter(B0015) E.coli

    • Digestion:[ 37oCRBS+mGFP+Ter(B0015)]XP

    • Single colony isolation from 37oCRBS+mGFP+Ter(B0015) LB-A plate, and cultivation in liquid LB-A.

    11

    • PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)]

    • Transformation of mRFP on LB-K plate.

    12

    • Single colony isolation from mRFP LB-K plate, and cultivation in liquid LB-K

    13

    • Ligation: insert[PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1C3]EP and insert[TetR]ES+[B0015]XP/vector[PSB1C3]EP.

    • Mini-prep of cultivated mRFP E.coli

    • Digestion:[mRFP]ES & [Ter(J61048)]XP & [37OCRBS+mGFP]XP

    • Ligation: insert 37oCRBS+LuxR[ES]&37oCRBS+mGFP[XP] /vector pSB1A3[EP]

    • Transformation of 37oCRBS +LuxR+37oCRBS+mGFP on LB-A plate & mRFP +Ter(J61048) on LB-C plate& Ter(J61048) on LB-A plate.

    14

    • Transformation of PompC+B0034+LacI+J61048+PSB1C3 and TetR+B0015+PSB1C3

    • Cultivation of B0032+ho1 with liquid LB-A. PCR of insert fragment[Ter(B0015)]

    • Single colony isolation from Ter(J61048) LB-A plate, and cultivation in liquid LB-A & 37OCRBS+mGFP on LB-K plate, and cultivation in liquid LB-K

    • Transformation of 37oCRBS +LuxR+37oCRBS+mGFP on LB-A plate & mRFP +Ter(J61048) on LB-C plate

    • Mini-prep of cultivated Ter (J61048)& 37oCRBS+mGFP E.coli

    15

    • Digestion:[B0032]XP

    • Transformation of PompC+B0034+LacI+J61048 +PSB1C3and TetR+B0015 +PSB1C3 again

    • Ligation: insert[PompC]ES+[B0032]XP/vector[PSB1K3]EP, then transformation of this part

    • Mini-prep of B0032+ho1. PCR of insert fragment[37oCRBS + LuxR + 37oCRBS+mGFP]&[mRFP+Ter(J61048)].

    9

    • Mini-prep from cultivated PompC+B0034+LacI+J61048 Eco.li&PSB1C3 E.coli

    • digestion : [PompC+B0034+LacI+J61048]ES&[PSB1C3]EP

    • Electrophoresis of [PompC+B0034+LacI+J61048] ES& [LacI+J61048]ES&[PSB1C3]EP.

    • Single colony isolation from 37oCRBS + mGFP+Ter(B0015)

    10

    • Mini-prep of cultivated 37oCRBS+mGFP+Ter(B0015) E.coli

    • Digestion:[ 37oCRBS+mGFP+Ter(B0015)]XP

    • Single colony isolation from 37oCRBS+mGFP+Ter(B0015) LB-A plate, and cultivation in liquid LB-A.

    11

    • PCR of insert fragment[37oCRBS+mGFP+Ter(B0015)]

    • Transformation of mRFP on LB-K plate.

    12

    • Single colony isolation from mRFP LB-K plate, and cultivation in liquid LB-K

    13

    • Ligation: insert[PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1C3]EP and insert[TetR]ES+[B0015]XP/vector[PSB1C3]EP.

    • Mini-prep of cultivated mRFP E.coli

    • Digestion:[mRFP]ES & [Ter(J61048)]XP & [37OCRBS+mGFP]XP

    • Ligation: insert 37oCRBS+LuxR[ES]&37oCRBS+mGFP[XP] /vector pSB1A3[EP]

    • Transformation of 37oCRBS +LuxR+37oCRBS+mGFP on LB-A plate & mRFP +Ter(J61048) on LB-C plate& Ter(J61048) on LB-A plate.

    14

    • Transformation of PompC+B0034+LacI+J61048+PSB1C3 and TetR+B0015+PSB1C3

    • Cultivation of B0032+ho1 with liquid LB-A. PCR of insert fragment[Ter(B0015)]

    • Single colony isolation from Ter(J61048) LB-A plate, and cultivation in liquid LB-A & 37OCRBS+mGFP on LB-K plate, and cultivation in liquid LB-K

    • Transformation of 37oCRBS +LuxR+37oCRBS+mGFP on LB-A plate & mRFP +Ter(J61048) on LB-C plate

    • Mini-prep of cultivated Ter (J61048)& 37oCRBS+mGFP E.coli

    15

    • Digestion:[B0032]XP

    • Transformation of PompC+B0034+LacI+J61048 +PSB1C3and TetR+B0015 +PSB1C3 again

    • Ligation: insert[PompC]ES+[B0032]XP/vector[PSB1K3]EP, then transformation of this part

    • Mini-prep of B0032+ho1. PCR of insert fragment[37oCRBS + LuxR + 37oCRBS+mGFP]&[mRFP+Ter(J61048)].

    16

    • PCR + single colony isolation of PompC+B003+LacI+J61048 Cr E.coli&TetR+B0015 Cr E.coli&PompC+B0032 Kr E.coli, then Electrophoresis of these parts

    17

    • Mini-prep of cultivated PompC+B0034 E.coli and TetR+B0015 Cr E.coli

    18

    19

    20

    • Ligation: Pcons+RBS+pcyA&B0032+ho1&pSB1A3.

    21

    • Ligation: insert[PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1A3]EP

    • Transformation of Pcons+RBS+pcyA+B0032+ho1 on LB-A plate. PCR of insert fragment[37oCRBS + LuxR + 37oCRBS+mGFP]

    • Digestion:[pSB1C3]EP&[mRFP]ES&[Ter(J61048)]XP

    • Ligation: insert mRFP[ES]&Ter(J61048)[XP] /vector pSB1C3[EP]

    • Transformation of mRFP + Ter (J61048) on LB-C plate

    • Single colony isolation from 37oCRBS + LuxR +37oCRBS+mGFP LB-A plate, and cultivation in liquid LB-A.

    22

    • Transformation of PompC+B0034+LacI+J61048+PSB1A3 but there is no colony appearing. PCR of insert fragment[mRFP+Ter(J61048)]

    • Mini-prep of cultivated 37o CRBS + LuxR + 37oCRBS+mGFP E.coli

    • Digestion:[ 37o CRBS + LuxR + 37oCRBS+mGFP]ES

    23

    • Digestion:[PompC+B0032]ES and [TetR+B0015]XP,[LacI+J61048]XP

    • Ligation: insert [PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1C3]EP

    • Transformation of PompC+B0034+LacI+J61048 +PSB1C3

    • Electrophoresis of [PompC+B0032]ES & [TetR+B0015]XP

    • Digestion again:[PompC+B0032]ES&[LacI+J61048]XP&[LacI+B0015]XP

    • Transformation of mRFP + Ter (J61048) on LB-C plate

    24

    • PCR and single colony isolation of PompC+B0034+LacI+J61048+PSB1C3 E.coli

    • Electrophoresis of [B0032]dig ES&[TetR+B0015]dig XP&[LacI+J61048]dig XP&PCR product [PompC+B0034+LacI+J61048] ,[TetR+B0015] OK

    • Decide to determine the sequence of [TetR+B0015] OK.

    • Single colony isolation from 6/21 plate.

    • Cultivation of Pcons+ RBS+pcyA+B0032+ho1 with liquid LB-A.

    • Electrophoresis of Pcons+RBS+PcyA+B0032+ho1.

    • PCR of insert fragment[mRFP+Ter(J61048)].

    25

    • Ligation:insert[PompC+B0034]ES+[LacI+J61048]XP&[PompC]ES+[B0032]XP/vector[PSB1C3]EP and [PompC+B0034]ES+[LacI+J61048]XP/vector[PSB1A3]EP

    • Transformation of the ligation product made today

    • PCR of Pcons+RBS+pcyA+B0032+ho1.

    • Cultivation of Pcons+RBS+pcyA+B0032+ho1 with liquid LB-A

    • Electrophoresis of Pcons+RBS+pcyA+B0032+ho1.

    • Mini-prep of Pcons+RBS+pcyA+B0032+ho1.

    • Ligation: insert mRFP[ES]&Ter(J61048)[XP]/vector pSB1C3[EP]

    26

    • PCR and single colony isolation of PompC+B0034+LacI+J61048+PSB1A3& PompC+B0034+LacI+J61048+PSB1C3&PompC+B0032+PSB1C3 E.coli

    • Electrophoresis of the PCR product made today

    • Electrophoresis of Pcons+RBS+pcyA+B0032+ho1──NOT OK

    • Ligation: Pcons+RBS+pcyA&B0032+ho1

    • Transformation of Pcons+RBS+pcyA+B0032+ho1. Transformation of mRFP + Ter (J61048) on LB-C plate

    27

    • Cultivation of PompC+B0034+LacI+J61048+PSB1A3 E.coli in liquid LB-A tubes

    • PCR of Pcons+RBS+pcyA+B0032+ho1

    • Cultivation of Pcons+RBS+pcyA+B0032+ho1 with liquid LB-A

    • Electrophoresis of Pcons+RBS+pcyA+B0032+ho1──NOT OK

    • Transformation of mRFP + Ter (J61048) on LB-C plate

    28

    • PCR of Pcons+RBS+pcyA+B0032+ho1

    • Electrophoresis of Pcons+RBS+pcAa+B0032+ho1──NOT OK

    • Ligation: insert mRFP[ES]&Ter(J61048)[XP] /vector pSB1C3[EP]

    29

    • Cultivation of PompC+B0034+LacI+J61048+PSB1A3 E.coli in liquid LB-A tubes again.

    • Ligation: Pcons+RBS+pcyA&B0032+ho1&pSB1A3

    • Transformation of ligated Pcons+RBS+pcyA+B0032+ho1

    • Single colony isolation from 6/21 Pcons+RBS+pcyA+B0032+ho1 LB plate

    • PCR of 6/21 Pcons+RBS+pcyA+B0032+ho1

    • Electrophoresis of 621 Pcons+RBS+pcyA+B0032+ho1─NOT OK

    • Transformation of mRFP+Ter(J61048) on LB-C plate

    • Single colony isolation from 37oCRBS +LuxR +37oCRBS+mGFP LB-A plate, and cultivation in liquid LB-A

    30

    • Mini-prep of cultivated PompC+B0034+LacI+J61048+PSB1A3 E.coli and then digest with EcoR1 and SpeI:[ PompC+B0034+LacI+J61048]dig ES

    • Electrophoresis of [PompC+B0034+LacI+J61048+PSB1A3] mini and [ PompC+B0034+LacI+J61048]dig ES OK

    • Decide to determine sequence of[ PompC+B0034+LacI+J61048]

    • Cultivation of 6/29 Pcons+RBS+PcyA+B0032+ho1 LB plate with liquid LB-A

    • PCR of 6/29 Pcons+RBS+PcyA+B0032+ho1

    • Cultivation of 6/29 re-ligated Pcons+RBS+PcyA+B0032+ho1

    • PCR of 6/29 re-ligated Pcons+RBS+pcyA+B0032+ho1

    • Electrophoresis of all above─NOT OK

    July 2013

    Sunday

    Monday

    Tuesday

    Wednesday

    Thursday

    Friday

    Saturday

    1

    • PCR + single colony isolation of PompC+B003+LacI+J61048 Cr E.coli&TetR+B0015 Cr E.coli&PompC+B0032 Kr E.coli, then Electrophoresis of these parts

    2

    • DNA sequencing OK

    3

    4

    5

    6

    7

    • digestion : pLac+B0034+zif268+AlsS (pm) [ES] & B0034+ PBSII+ilvC (pm) [XP]

    8

    9

    • Testing the temperature of the point mutation between of HivC & ilvD by the m.p 50℃ of PCR

    • digestion : pSB1K3 [EP]

    • ligation : insert pLac+B0034+zif268+AlsS (pm) [ES] & B0034+ PBSII+ilvC (pm) [XP]/vector pSB1K3 [EP]

    • transformation of pLac+B0034+zif268+AlsS+ B0034+ PBSII+ilvC (pm) and cultivation on LB-K plate

    10

    • Testing the temperature of the point mutation between of HivC & ilvD by the m.p 48℃ of PCR

    • Digestion : B0034+ HivC+ilvD+37℃ RBS [DPn1]

    • PCR of insert fragment [pLac+B0034+zif268+AlsS+ B0034+ PBSII+ilvC] (pm) OK

    • SCI from pLac+B0034+zif268+AlsS+ B0034+ PBSII+ilvC (pm) LB-K plate, and cultivation in liquid LB-K

    11

    • mini-prep of cultivated pLac+B0034+zif268+AlsS+ B0034+ PBSII+ilvC (pm) E. coli

    • DNA sequencing OK

    • culture condition test: activation DH5αovernight

    12

    • transfer to new medium(1/100), OD0.2 start counting culture time

    • transfer to 30゜C and 27゜C

    13

    • transfer to 30゜C and 27゜C

    14

    15

    • transformation of B0034+ HivC+ ilvD+37℃ RBS (pm),and cultivation on LB-A plate

    16

    • PCR of insert fragment [B0034+ HivC+ilvD+37℃ RBS](pm)-----NOT OK

    17

    • Point mutation by PCR of insert fragment [B0034+ HivC+ilvD+37℃ RBS]

    • Digestion: B0034+ HivC+ilvD+37℃ RBS [DPn1]

    18

    19

    • transformation of B0034+ HivC+ ilvD+37℃ RBS (pm),and cultivation on LB-A,K,C plate (A sucessful)

    20

    • PCR of insert fragment [B0034+ HivC+ilvD+37℃ RBS](pm) ---- OK

    • Single colony isolation from B0034+ HivC+ilvD+37℃ RBS (pm) LB-A plate, and cultivation in liquid LB-A

    21

    • mini-prep of cultivated B0034+ HivC+ilvD+37℃ RBS (pm) E. coli

    • Digestion: pLac+B0034+zif268+AlsS+ B0034+ PBSII+ilvC (pm)(G1) [ES] & B0034+ HivC+ilvD+37℃ RBS (pm)(G2) [XP]

    • ligation : insert G1[ES] & G2[XP] vector pSB1C3[EP]

    22

    • transformation of G1+G2,and cultivation on LB- C plate failed

    23

    • ligation : insert G1[ES] & G2[XP]/vector pSB1C3[EP]

    24

    • transformation of G1+G2,and cultivation on LB- C plate

    • Digestion: G2’(B0034+HIVC+ilvD) [XP]

    • ligation : insert G1[ES] & G2‘[XP]/vector pSB1C3[EP]

    25

    • PCR of insert fragment [kivD+B0015]

    • transformation of G1+G2’, and cultivation on LB- C plate failed

    26

    • Single colony isolation from kivD+B0015 LB-C plate, and cultivation in liquid LB-C

    • sample and do GC

    27

    • mini-prep of cultivated kivD+B0015 E. coli

    28

    29

    • Point mutation by PCR of insert fragment [B0034+ HivC+ilvD(G2’)]

    • Digestion: G2’ [DPn1]

    30

    • transformation of G2’, and cultivation on LB- A plate

    31

    • Digestion: kivD+B0015 [EP] (checking bp----failed)

    • Single colony isolation from G2’ (pm) LB-A plate, and cultivation in liquid LB-A

    August 2013

    Sunday

    Monday

    Tuesday

    Wednesday

    Thursday

    Friday

    Saturday

    1

    • mini-prep of cultivated G2’ E. coli

    2

    3

    4

    5

    6

    7

    8

    9

    • Digestion: G1 [ES] & G2‘ [XP] & pSB1C3 [EP]

    • ligation : insert G1[ES] & G2‘[XP]/vector pSB1C3[EP]

    10

    • strain test: activation of different strains overnight

    11

    • transfer to new medium, OD0.2 IPTGinduction, culture in 37゜C

    12

    • Single colony isolation from G1+G2’ LB-C plate, and cultivation in liquid LB-C

    • transfer to 27゜C

    13

    • mini-prep of cultivated G1+G2’ E. coli

    14

    15

    • Digestion: Ptet+B0032[ES] & GliI+KivD[XP]

    • sample and do GC

    16

    • ligation : insert Ptet+B0032[ES] & GliI+KivD[XP]/vector pSB1K3[EP]

    • Single colony isolation from G1+G2 LB-C plate, and cultivation in liquid LB-C

    17

    • transformation of Ptet+B0032+ GliI+KivD, and cultivation on LB- K plate

    • mini-prep of cultivated G1+G2 E. coli

    18

    • PCR of insert fragment [Ptet+B0032+ GliI+KivD]-----OK

    • DNA sequencing------NOT OK

    19

    20

    21

    22

    23

    24

    • carbon source test: activation DH5α overnight

    25

    • transfer to new medium(1/100), OD0.2 start counting culture time

    26

    • culturing for 72hours, inject feeding solution per 24hours

    27

    • culturing for 72hours, inject feeding solution per 24hours

    28

    • transformation of DNA program, and cultivation on LB- A plate

    • culturing for 72hours, inject feeding solution per 24hours

    29

    • ligation : insert Ptet+B0032[ES] & GliI+KivD[XP]/vector pSB1K3[EP]

    • Single colony isolation from DNA program LB-C plate, and cultivation in liquid LB-C

    • sample & do GC

    30

    • transformation of Ptet+B0032+ GliI+KivD, and cultivation on LB- K plate

    • mini-prep of cultivated DNA program E. coli

    • Do GC

    31

    • PCR of insert fragment [Ptet+B0032+ GliI+KivD] OK

    • DNA sequencing NOT OK

    September 2013

    Sunday

    Monday

    Tuesday

    Wednesday

    Thursday

    Friday

    Saturday

    1

    • ligation : insert Zif268 [ES] & AlsS [XP]/Vector pSB1K3 [EP]

    2

    • transformation of Zif268+AlsS+pSB1K3 and cultivation on LB-K plate

    3

    4

    • digestion : [pSB1K3] EP

    • digestion : [pSB1K3] EP

    5

    6

    7

    • PCR of insert fragment [Zif268+AlsS] OK

    • DNA Sequencing OK

    • ligation : point mutation HivC

    • TA cloning : point mutation HivC

    8

    9

    • Single colony isolation from Zif268+AlsS LB-K plate, and cultivation of in liquid LB-K

    10

    • mini-prep of cultivated Zif268+ AlsS E. coli

    • transformation of point mutation HivC and cultivation on LB-A plate

    • transformation of B0034 and cultivation on LB-A plate

    11

    12

    • Single colony isolation from HivC LB-A plate, and in liquid LB-A

    • Single colony isolation from B0034 LB-A plate, and in liquid LB-A

    13

    • mini-prep of cultivated point mutation HivC E. coli & B0034 E. coli

    14

    • digestion : Zif268+AlsS [XP]

    15

    16

    17

    • Single colony isolation from ilvD LB-A plate, and cultivation in liquid LB-A

    18

    • mini-prep of cultivated ilvD E. coli

    • digestion : ilvD [EP] & pSB1K3 [EP]

    • transformation of 37℃ RBS and cultivation on LB-C plate

    19

    • Single colony isolation from HivC LB-A plate, and cultivation in liquid LB-A

    20

    • digestion : B0034 [SP]

    • gel extraction

    • ligation :insert Zif268+AlsS [XP]/Vector B0034 [SP]

    • mini-prep of cultivated Hivc E. coli

    21

    • transformation of B0034+Zif268+AlsS and cultivation on LB-A plate

    • transformation of HivC and cultivation on LB-A plate

    22

    • PCR of insert fragment [B0034+Zif268+AlsS] OK

    • DNA Sequencing NOT OK

    • digestion : B0034 [SP] & Zif268+AlsS [XP]

    • gel extraction

    • ligation :insert Zif268+AlsS [XP]/Vector B0034+pSB1A3 [SP]

    • transformation of B0034+Zif268+AlsS and cultivation on LB-A plate

    • Single colony isolation from B0034 LB-A plate, and cultivation in liquid LB-A

    • SCI from ilvD LB-K plate, and cultivation in liquid LB-K

    23

    • PCR of insert fragment [B0034+Zif268+AlsS] NOT OK

    • transformation of B0034+Zif268+AlsS and cultivation on LB-A plate

    • mini-prep of cultivated B0034 and ilvD E. coli

    • Single colony isolation from HivC LB-A plate, and cultivation in liquid LB-A

    24

    • mini-prep of cultivated HivC E. coli

    • digestion : B0034 [SP] & ilvD [XP]

    25

    • digestion : pSB1C3 [EP] & HivC [ES] & HivC [SP]

    • ligation :insert HivC [ES] & ilvD [XP]/Vector pSB1C3 [EP]

    • ligation :insert ilvD [XP]/Vector HivC+pSB1A3 [SP]

    26

    • transformation of HivC+ilvD and cultivation on LB-A & LB-C plate

    27

    • PCR of insert fragment [B0034+Zif268+AlsS] NOT OK

    28

    • transformation of pSB1A3 & pSB1C3 and cultivation on LB-A & LB-C plate

    • PCR of insert fragment [HivC+ilvD] OK

    • DNA sequencing NOT OK

    29

    • Single colony isolation from pSB1C3 LB-C plate, and cultivation in liquid LB-C

    30

    • mini-prep of cultivated & pSB1C3 E. coli

    Retrieved from "http://2013.igem.org/Team:NCTU_Formosa/notes"