Difference between revisions of "Haynes Lab:Notebook/Engineering PC-TFs/2013/04/01"

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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Engineering PC-TFs</span>
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|style="background-color: #FFCC00"|<br>[[Image:Hayneslab3.gif|200px|center]][[Image:Asu logo 3.gif|200px|center]]<span style="font-size:22px;"><br></span>
|style="background-color: #F2F2F2" align="center"|<html><img src="/images/9/94/Report.png" border="0" /></html> [[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>[[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]]<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>}}
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|style="background-color: #800000" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}}
 
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==Summary==
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==4/2/13==
 
*1. Kozak,V0120 + BL01
 
*1. Kozak,V0120 + BL01
 
*2. Kozak,V0120 + BL05
 
*2. Kozak,V0120 + BL05

Latest revision as of 21:35, 26 September 2017


Hayneslab3.gif
Asu logo 3.gif

Report.pngMain project page
Resultset previous.pngPrevious entry      Next entryResultset next.png

4/2/13

  • 1. Kozak,V0120 + BL01
  • 2. Kozak,V0120 + BL05
  • 3. Kozak,V0120 + BL09
  • 4. Kozak,V0120 + BL12
# Biobrick ng/µL
1 hPCD : mCh : SP1AB (BL01) 18.834
2 fshPCD : mCh : SP1AB (BL05) 14.471
3 flyPCD : mCh : SP1A (BL09) 7.302
4 hPCD : BL01 (BL12) 10.84








µL of insert = (length insert/lengthvector *2*25 ng vector/concentration insert)
1 2 3 4 5
DNA Insert 1.4 1.8 2.0 2.5 --
DNA Vector (Kozak) 0.6 0.6 0.6 0.6 0.6
T4 Ligase 1 1 1 1 1
Lign Buffer (2x) 5 5 5 5 5
dH2O 2.0 1.6 1.4 .9 3.7
10 µL 10 µL 10 µL 10 µL 10 µL
  • In a .5 mL tube, pipette the ligation buffer first.
  • Then water, DNA insert, vector, and T4 ligase.
  • Once completed, allow them to incubate for 10 minutes.
  • Meanwhile, get ice bucket and allow competent E. coli cells to thaw for five minutes.
  • Then follow transformation process:

Transformation Process

  • Warm five 100 μg/mL Amp agar plates at 37 °C
  • Thaw fresh tube of DH5α Turbo cells on ice
  • Add 30 μL of DH5α Turbo cells to DNA + dH2O
  • Include #5 tube, water only, no DNA (negative control)
  • Incubate cells + DNA on ice for 10 min.
  • Label pre-warmed plates
  • Transfer cells + DNA onto agar
  • Add 10 - 15 sterile glass beads, shake, discard beads
  • Incubate plates at 37 °C overnight
Ligation attempt 4/2/2013
  • No colonies on any of the plates. Try again.