Team:Lethbridge/Notebook/Lab Work/October

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<b>Objective:</b> PCR of xylE for conformation of transformation.
<b>Objective:</b> PCR of xylE for conformation of transformation.
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<table border ="3">
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<tr><td><b>Component</b><td><b>1X(&micro;L)</b><td><b>Master Mix(x24)(&micro;L)</b>
 +
<tr><td>Milli-Q H<sub>2</sub>O<td>13.8<td>331.2
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<tr><td>10x Pfu Buffer with MgSO<sub>4</sub><td>2<td>48
 +
<tr><td>dNTPs<td>1<td>24
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<tr><td>Forward VF2 Primer<td>1<td>24
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<tr><td>Reverse VR Primer<td>1<td>24
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<tr><td>Template DNA<td>1<td>
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<tr><td>Pfu polymerase<td>0.2<td>4.8
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</table><br>
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 +
Ran with PFV program on thermocycler with extension time of 3 minutes.
==<font color="white">October 2, 2010==
==<font color="white">October 2, 2010==

Revision as of 18:16, 23 October 2010



Feel free to look around our notebook!

Here you can check out the work we have done in the lab, click on a month to take a look!

Contents

October

October 1, 2010

KG

Objective: PCR of xylE for conformation of transformation.

Component1X(µL)Master Mix(x24)(µL)
Milli-Q H2O13.8331.2
10x Pfu Buffer with MgSO4248
dNTPs124
Forward VF2 Primer124
Reverse VR Primer124
Template DNA1
Pfu polymerase0.24.8

Ran with PFV program on thermocycler with extension time of 3 minutes.

October 2, 2010

ADS

Objective: Purify plasmid DNA from transformed Mr. GENE synthesized DNA

  • <partinfo>K331022</partinfo>
  • <partinfo>K331023</partinfo>
  • <partinfo>K331024</partinfo>
  • <partinfo>K331025</partinfo>

Method: Use Qiagen miniprep method with BioBasic EZ-10 Spin columns.

Objective: Create 20 new biobricks using recently arrived synthesized DNA and terminator <partinfo>B0015</partinfo>, pLac <partinfo>R0010</partinfo>, and pTet <partinfo>R0040</partinfo> to create the following new biobricks:

  • 1)R0010 + K331022 to obtain <partinfo>K331026</partinfo>
  • 2)R0040 + K331022 to obtain <partinfo>K331030</partinfo>
  • 3)K331022 + B0015 to obtain <partinfo>K331034</partinfo>
  • 4)K331022 + K331024 to obtain (NAME BIOBRICK!!!!)
  • 5)R0010 + K331023 to obtain <partinfo>K331027</partinfo>
  • 6)R0040 + K331023 to obtain <partinfo>K331031</partinfo>
  • 7)K331023 + B0015 to obtain <partinfo>K331035</partinfo>
  • 8)K331023 + K331025 to obtain (NAME BIOBRICK!!!!)
  • 9)R0010 + K331024 to obtain <partinfo>K331028</partinfo>
  • 10)R0040 + K331024 to obtain <partinfo>K331032</partinfo>
  • 11)K331024 + B0015 to obtain <partinfo>K331036</partinfo>
  • 12)K331024 + K331022 to obtain (NAME BIOBRICK!!!!)
  • 13)R0010 + K331025 to obtain <partinfo>K331029</partinfo>
  • 14)R0040 + K331025 to obtain <partinfo>K331033</partinfo>
  • 15)K331025 + B0015 to obtain <partinfo>K331037</partinfo>
  • 16)K331025 + K331023 to obtain (NAME BIOBRICK!!!!)
  • 17)K331022 to obtain <partinfo>K331022</partinfo>
  • 18)K331023 to obtain <partinfo>K331023</partinfo>
  • 19)K331024 to obtain <partinfo>K331024</partinfo>
  • 20)K331025 to obtain <partinfo>K331025</partinfo>

All parts will be inserted into pSB1C3.
Method: Use BioBrick Assembly Method
Results: Obtained RESULTS!!! positive colonies.

October 5, 2010

ADS

Objective: Purify plasmid DNA from successfully assembled biobricks.
Method: Use Qiagen miniprep method with BioBasic EZ-10 columns.
Will send these minipreps for sequencing at Macrogen.




October 16, 2010

JV

Objective: Separate different fractions of the cytosol of cells containing catechol 2,3-dioxygenase
Method:

  • Grow DH5α cells in 500mL of LB media with Amp+.
  • Optical density of cell suspension (measured at 600nm): 4.55
  • Spun cells down: 10 minutes, 4oC, at 5000RPM
  • Cell pellet weight is 3.03g
  • Flash froze cells and stored at -80oC

Plasmid DNA was also isolated using QIAGEN spin column protocol. DNA was eluted to 60µL.

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