Team:HokkaidoU Japan/Notebook/plastic protocols

From 2012.igem.org

(Difference between revisions)
(→Polymer producing media)
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|10ml
|10ml
|-
|-
-
|50% Glucose
+
|●50% Glucose
|800ul
|800ul
|-
|-
-
|1M pantothenic acid Ca
+
|●1M pantothenic acid Ca
|200ul
|200ul
|-
|-
Line 28: Line 28:
Put 1.5ml each into the test tube.
Put 1.5ml each into the test tube.
-
polymer producing media (LB:2%Glc+ 10mM pantothenic acid Ca + Amp100mg/l) 20ml
+
 
 +
●50%gulcose
{|class="hokkaidou-table-digestion"
{|class="hokkaidou-table-digestion"
|-
|-
-
|2×LB
+
|ROwater
-
|10ml
+
|7ml
|-
|-
-
|50% Glucose
+
|Glucose
-
|800ul
+
|10g  Heat until it melts.
|-
|-
-
|1M pantothenic acid Ca
+
|ROwater
-
|200ul
+
|up to 20ml
|-
|-
-
|Amp(100mg/ml)
+
|}
-
|20ul
+
 
 +
Filter sterilize.
 +
 
 +
 
 +
●1M pantothenic acid Ca
 +
{|class="hokkaidou-table-digestion"
|-
|-
|ROwater
|ROwater
-
|8.98ml
+
|7ml
 +
|-
 +
|pantothenic acid Ca
 +
|4.77g  Heat　and stir until it melts.
 +
|-
 +
|ROwater
 +
|up to 10ml
|-
|-
|}
|}
-
</p>
 
 +
Filter sterilize.
 +
 +
 +
 +
</p>
===Polymer extraction and purification===
===Polymer extraction and purification===

Revision as of 09:32, 6 August 2012

Polymer producing media

polymer producing media (LB:2%Glc+ 10mM pantothenic acid Ca + Amp100mg/l) 20ml

2×LB 10ml
●50% Glucose 800ul
●1M pantothenic acid Ca 200ul
Amp(100mg/ml) 20ul
ROwater 8.98ml

Put 1.5ml each into the test tube.


●50%gulcose

ROwater 7ml
Glucose 10g Heat until it melts.
ROwater up to 20ml

Filter sterilize.


●1M pantothenic acid Ca

ROwater 7ml
pantothenic acid Ca 4.77g Heat　and stir until it melts.
ROwater up to 10ml


Filter sterilize.


Polymer extraction and purification

  1. Move dried up bacteria into test tube.
  2. Break up them to separate and add 10ml chloroform.
  3. Incubate for 48hour at 60C.
  4. Make it filtered through PTFE and move it into another test tube.
  5. Volatilize organic solvent by exposing air and separate polymer.
  6. Add 5ml hexane and voltex for a minute. After centrifuging (1,500rpm, 10min), remove the clear layer.
  7. Volatilize chloroform by exposing air again.
  8. Add 5ml MtOH, voltex for a minute, centrifuge (1,500rpm, 10min), and remove the clear layer.



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