Haynes Lab:Notebook/Engineering PC-TFs/2013/01/25

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# pSB1A3, gg0001, gg0002
# pSB1A3, gg0001, gg0002
-
# template, primer 1, primer 2
+
# hPCD, gg0003, gg0004
-
# template, primer 1, primer 2
+
# BL02, gg0005, gg0007
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# template, primer 1, primer 2
+
# BL03, gg0005, gg0008
 +
# BL04, gg0005, gg0009
 +
# fshPCD, gg0010, gg0012
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| bgcolor=#cfcfcf | Reagent
| bgcolor=#cfcfcf | Reagent
| bgcolor=#cfcfcf | Volume  
| bgcolor=#cfcfcf | Volume  
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| rowspan="7" | <u>Expected:</u><br>1. pSB1A3 = ~2000<br>2. hPCD = 186<br>3. BL01 = 2520
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| rowspan="7" | <u>Expected:</u><br>1. pSB1A3 = ~2000<br>2. hPCD = 186<br>3. BL02 = 1431<br> 4. BL03 = 1731<br> 5. BL04 = 1686<br> 6.fshPCD = 186<br>
| rowspan="7" | [[Image:gg_gel1-25-2013.jpg|200px|PCR results]]<br>10 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
| rowspan="7" | [[Image:gg_gel1-25-2013.jpg|200px|PCR results]]<br>10 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
|-
|-
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Purification
+
Purification (Completed on 1-25-2013)
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* Clean with Zymo DNA c&c; elute w/ 20 μL dH<sub>2</sub>O
+
* Clean with Zymo DNA c&c; elute w/ 20 μL dH<sub>2</sub>O  
-
* Measure [DNA]
+
 
 +
* Measure [DNA] (1-29-2013)
{| {{table}} border="1" cellspacing="3" <!-- Digest check rxn. table -->
{| {{table}} border="1" cellspacing="3" <!-- Digest check rxn. table -->
|- valign="top"
|- valign="top"

Revision as of 01:35, 28 January 2013

Engineering PC-TFs Main project page
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01/25/13

  • Type IIS assembly - day 1



PCR

  • Resuspend new primers to a final concentration of 100 μM
  • Make 10 μM dilutions in separate tubes
  • All of the reactions are as follows:
  1. pSB1A3, gg0001, gg0002
  2. hPCD, gg0003, gg0004
  3. BL02, gg0005, gg0007
  4. BL03, gg0005, gg0008
  5. BL04, gg0005, gg0009
  6. fshPCD, gg0010, gg0012


Reagent Volume Expected:
1. pSB1A3 = ~2000
2. hPCD = 186
3. BL02 = 1431
4. BL03 = 1731
5. BL04 = 1686
6.fshPCD = 186
PCR results
10 μL/lane; 1% agarose; Ladder
DNA(plasmid) 0.2 μL
primer 1 1.0
primer 2 1.0
2x GoTaq 25.0
dH2O 22.8
  50.0 μL
  • 95°C, 3 min.
  • [95°C, 30 sec.; 57°C, 30 sec.; 72°C, 1 min.] x35
  • 72°C, 3 min.
  • 4°C, ∞


Purification (Completed on 1-25-2013)

  • Clean with Zymo DNA c&c; elute w/ 20 μL dH2O
  • Measure [DNA] (1-29-2013)
Sample OD 260 260/280 ng/μL
1. pSB1A3 ### ### ###
2. ### ### ### ###
3. ### ### ### ###



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