Difference between revisions of "IGEM:University of East Anglia (UEA), Norwich, UK/2009/Notebook/NRP-UEA-Norwich iGEM/2014/07/10"

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(Autocreate 2014/07/10 Entry for IGEM:University_of_East_Anglia_(UEA),_Norwich,_UK/2009/Notebook/NRP-UEA-Norwich_iGEM)
 
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|style="background-color: #EEE"|[[Image:igem-logo-150px.png|150px]]<span style="font-size:22px;"> iGEM Project name 1</span>
 
|style="background-color: #EEE"|[[Image:igem-logo-150px.png|150px]]<span style="font-size:22px;"> iGEM Project name 1</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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==Entry title==
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==Week 4 Day 4==
* Insert your content here.
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TSL
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----
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- Agrobacterium-mediated transient expression in leaves of ''N. benthamiana'' PROTOCOL:
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1. Single colony of each constructs 2,3,4,10,11 grown overnight at 28 °C in 5 ml media containing chloramphenicol.
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2. Pellet by centrifugation at 4000 rpm for 15 min.
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3. Resuspend in 10 mM MES, 10 mM MgCl2 and 150 μM Acetosyringone.
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4. 1:100 dilution to give a final OD of 0.1-0.5
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5. Incubate at RT for 4 hours.
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6. Create a small puncture using a needle on the underside of ''N. benthamiana'' leaves of plants 6-8 weeks old. Using a blunt syringe infiltrate the agro constructs into the leaf puncture. Each construct is infiltrated into 4 sections on one leaf. 2 constructs per plant.
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7. Store plants in controlled environment at 19-23 °C for 3 days.  
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-Ligation reaction to join PSB1C3 and GG compatible ends
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•PCR program?
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•Transfored into E.coli
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•Spread plated onto CAM plates
  
  

Latest revision as of 00:06, 27 September 2017

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Week 4 Day 4

TSL


- Agrobacterium-mediated transient expression in leaves of N. benthamiana PROTOCOL:

1. Single colony of each constructs 2,3,4,10,11 grown overnight at 28 °C in 5 ml media containing chloramphenicol.

2. Pellet by centrifugation at 4000 rpm for 15 min.

3. Resuspend in 10 mM MES, 10 mM MgCl2 and 150 μM Acetosyringone.

4. 1:100 dilution to give a final OD of 0.1-0.5

5. Incubate at RT for 4 hours.

6. Create a small puncture using a needle on the underside of N. benthamiana leaves of plants 6-8 weeks old. Using a blunt syringe infiltrate the agro constructs into the leaf puncture. Each construct is infiltrated into 4 sections on one leaf. 2 constructs per plant.

7. Store plants in controlled environment at 19-23 °C for 3 days.


-Ligation reaction to join PSB1C3 and GG compatible ends •PCR program? •Transfored into E.coli •Spread plated onto CAM plates