View source for IGEM:Missouri University of Science and Technology/2009/Notebook/Cyanobacteria Mediated Filtration of Coal Flue/2013/07/31

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|style="background-color: #EEE"|[[Image:igem-logo-150px.png|150px]]<span style="font-size:20px;"> Cyanobacteria Mediated Filtration of Coal Flue Gas</span>
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== <b>Date: 7/31/13</b> <b>People in lab: Emily Puleo, Kelsey Crossen</b>  ==

<b><span style="font-size:16px;">Title:</span></b> PCR of norCB, dilution of norCB primers

<b>Start Time:</b> 4:57 PM

<b>Purpose:</b> PCR norCB from P. aeruginosa, adding iGEM prefix and suffix; to prepare primers for future lab work

<b>Protocol:</b> LTM ed. 2 pg. 48 <b>Exceptions:</b> 1. Less than 0.1 final primer concentration. 

<b>Products:</b> 
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! Sample Label !! Description !! Source Label !! Quantity
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| 7/31/13 A norCB 1 || Amplified norCB, with prefix/suffix, 7.5uL template || P. aeruginosa 7/30 || 1
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| 7/31/13 A norCB 2 || Amplified norCB, with prefix/suffix, 5.63uL template || P. aeruginosa 7/30 || 1
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| 7/31/13 A norCB 3 || Amplified norCB, with prefix/suffix, 3.75uL template || P. aeruginosa 7/30 || 1 
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| 7/31/13 norCB F || Forward primer for norCB, 0.73 uM || norCB Forward Primer || 1
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| 7/31/13 norCB R || Reverse primer for norCB, 0.87 uM || norCB Reverse Primer || 1
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<b>Results:</b> Sample 3 had too little product and was discarded.

<b>Notes:</b> Different amounts of template used to find optimum amount needed.

Forward primer: (24.2*10^-9)/(0.33*10^-3) mol/L = 73 uM,  (73uM)(10uL)=(0.73uM)V2  V2 = 1000uL

Reverse primer: (28.7*10^-9)/(0.33*10^-3) mol/L = 87 uM,  (87uM)(10uL)=(0.87uM)V2  V2 = 1000uL

Used Dr. Shannon's thermocycler, copied "igem_pcr" program

<b>Stop Time:</b> 9:00 PM

<b>Next:</b> Run samples on a gel, extract the stronger band from between them, sequence the result.



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