Difference between revisions of "Team:Nagahama/Design"

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{{Nagahama}}
 
{{Nagahama}}
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= '''Protocols''' =
 +
Our Lab's Protocols
 +
 +
== Medium ==
 +
 +
=== LB medium (100 mL liquid) ===
 +
 +
1.Measure 1g Tripton
 +
 +
2.Measure 0.5g Yeast Extract
 +
 +
3.Measure 1g Nacl
 +
 +
4.Add 100mL H2O
 +
 +
5.autoclave(121℃ 20min)
 +
 +
=== 2×YT medium (100mL liquid)===
 +
 +
1.Measure 1.6g Tripton
 +
 +
2.Measure 1g Yeast Extract
 +
 +
3.Measure 0.5g Nacl
 +
 +
4.Add 100mL H2O
 +
 +
5.autoclave(121℃ 20min)
 +
 +
 +
== DNA work ==
 +
 +
 +
=== Agarose gel(100mL) ===
 +
 +
Method of Making 0.7% Agarose gel
 +
 +
1.Measure 0.7g Agarose
 +
 +
2.Add 100mL TAE buffer
 +
 +
3.Heat(till agarose melted)*We used a microwave oven.
 +
 +
4.Pur agarose into a gel maker
 +
 +
5.Set a comb
 +
 +
6.Wait till agarose curdles
 +
 +
7.Pull an comb
 +
 +
===  Genome DNA extraction ===
 +
↓Cultivate ''E. coli DH5α'' using LB medium 2ml×2 tubes O/N
 +
 +
↓Centrifuge culture 1.5ml  (13,000rpm　4℃　1min)
 +
 +
↓Remove the culture
 +
 +
↓+TNE buffer 1.0ml  (10mM Tris pH 8.0, 10 mM NaCl, 10 mM EDTA)
 +
 +
↓vortex
 +
 +
↓Centrifuge cell suspension  (13,000rpm　4℃　2min)
 +
 +
↓Remove  supernatant
 +
 +
↓+TNE buffer including 1% Triton X-100  270µl
 +
 +
↓Suspend cell gently
 +
 +
↓+5mg/ml Lysozyme solution 30µl (0.15g Lyzozyme + 30µl sterile water)
 +
 +
↓Reaction 37℃　30min
 +
 +
↓+ 20mg/ml Proteinase K (fin.con: 1mg/ml)    TaKaRa Code:9033
 +
 +
↓Reaction 65℃ 2h
 +
 +
↓+Phenol chloroform 300µl ...(1)
 +
 +
↓Mix the solution gently ...(2)
 +
 +
↓Centrifuge (13,000rpm　4℃　8min) ...(3)
 +
 +
↓Transfer only water layer to new 1.5ml tube ...(4)
 +
 +
↓Repeat (1)~(4) 2 times
 +
 +
↓+3M Sodium acetate 30µl
 +
 +
↓Mix gently
 +
 +
↓+99.5% EtOH 750µl
 +
 +
↓Mix gently
 +
 +
↓Centrifuge (13,000rpm　4℃　10min)
 +
 +
↓Remove supernatant
 +
 +
↓+70% EtOH 500µl
 +
 +
↓Centrifuge (13,000rpm　4℃　1min)
 +
 +
↓Dry up the pellet covering with aluminum foil at room temperature 30min
 +
 +
↓+TE buffer 50µl
 +
 +
↓Suspend DNA gently
 +
 +
[[File:GenomeDNA_DH5a2.png]]
 +
 +
=== Plasmids extraction ===
 +
 +
 +
 +
=== PCR ===
 +
 +
=== Transformation by heat shock===
 +
 +
== isoprenoid production ==
 +
 +
== geraniol tolerant assay ==
 +
 +
== GC ==
 +
 +
== GC-MS ==
 +
 +
 
 
<html>
 
<html>
  
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<h2>Design</h2>
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<h2>Experiments &amp; Protocols</h2>
  
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<p>
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<p>Describe the experiments, research and protocols you used in your iGEM project.</p>
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By talking about your design work on this page, there is one medal criterion that you can attempt to meet, and one award that you can apply for. If your team is going for a gold medal by building a functional prototype, you should tell us what you did on this page. If you are going for the <a href="https://2015.igem.org/Judging/Awards#SpecialPrizes">Applied Design award</a>, you should also complete this page and tell us what you did.  
+
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</p>
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<div class="highlightBox">
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<h5>What should this page contain?</h5>
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<h4>Note</h4>
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<ul>
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<p>In order to be considered for the <a href="https://2015.igem.org/Judging/Awards#SpecialPrizes">Best Applied Design award</a> and/or the <a href="https://2015.igem.org/Judging/Awards#Medals">functional prototype gold medal criterion</a>, you must fill out this page.</p>
+
<li> Protocols </li>
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</div>
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<li> Experiments </li>
 +
<li>Documentation of the development of your project </li>
 +
</ul>
  
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<p>This is a prize for the team that has developed a synthetic biology product to solve a real world problem in the most elegant way. The students will have considered how well the product addresses the problem versus other potential solutions, how the product integrates or disrupts other products and processes, and how its lifecycle can more broadly impact our lives and environments in positive and negative ways.</p>
 
  
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<p>
 
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If you are working on art and design as your main project, please join the art and design track. If you are integrating art and design into the core of your main project, please apply for the award by completing this page.
 
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</p>
 
  
 +
<h4>Inspiration</h4>
 +
<ul>
 +
<li><a href="https://2014.igem.org/Team:Colombia/Protocols">2014 Colombia </a></li>
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<li><a href="https://2014.igem.org/Team:Imperial/Protocols">2014 Imperial </a></li>
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<li><a href="https://2014.igem.org/Team:Caltech/Project/Experiments">2014 Caltech </a></li>
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</ul>
 
</div>
 
</div>
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</html>
 
</html>

Revision as of 05:39, 7 September 2015

Team Nagahama banner.jpg


Protocols

Our Lab's Protocols

Medium

LB medium (100 mL liquid)

1.Measure 1g Tripton

2.Measure 0.5g Yeast Extract

3.Measure 1g Nacl

4.Add 100mL H2O

5.autoclave(121℃ 20min)

2×YT medium (100mL liquid)

1.Measure 1.6g Tripton

2.Measure 1g Yeast Extract

3.Measure 0.5g Nacl

4.Add 100mL H2O

5.autoclave(121℃ 20min)


DNA work

Agarose gel(100mL)

Method of Making 0.7% Agarose gel

1.Measure 0.7g Agarose

2.Add 100mL TAE buffer

3.Heat(till agarose melted)*We used a microwave oven.

4.Pur agarose into a gel maker

5.Set a comb

6.Wait till agarose curdles

7.Pull an comb

Genome DNA extraction

↓Cultivate E. coli DH5α using LB medium 2ml×2 tubes O/N

↓Centrifuge culture 1.5ml (13,000rpm　4℃　1min)

↓Remove the culture

↓+TNE buffer 1.0ml (10mM Tris pH 8.0, 10 mM NaCl, 10 mM EDTA)

↓vortex

↓Centrifuge cell suspension (13,000rpm　4℃　2min)

↓Remove supernatant

↓+TNE buffer including 1% Triton X-100 270µl

↓Suspend cell gently

↓+5mg/ml Lysozyme solution 30µl (0.15g Lyzozyme + 30µl sterile water)

↓Reaction 37℃　30min

↓+ 20mg/ml Proteinase K (fin.con: 1mg/ml) TaKaRa Code:9033

↓Reaction 65℃ 2h

↓+Phenol chloroform 300µl ...(1)

↓Mix the solution gently ...(2)

↓Centrifuge (13,000rpm　4℃　8min) ...(3)

↓Transfer only water layer to new 1.5ml tube ...(4)

↓Repeat (1)~(4) 2 times

↓+3M Sodium acetate 30µl

↓Mix gently

↓+99.5% EtOH 750µl

↓Mix gently

↓Centrifuge (13,000rpm　4℃　10min)

↓Remove supernatant

↓+70% EtOH 500µl

↓Centrifuge (13,000rpm　4℃　1min)

↓Dry up the pellet covering with aluminum foil at room temperature 30min

↓+TE buffer 50µl

↓Suspend DNA gently

GenomeDNA DH5a2.png

Plasmids extraction

PCR

Transformation by heat shock

isoprenoid production

geraniol tolerant assay

GC

GC-MS

Experiments & Protocols

Describe the experiments, research and protocols you used in your iGEM project.

What should this page contain?
  • Protocols
  • Experiments
  • Documentation of the development of your project

Inspiration