Team:Tokyo Metropolitan/Project/Fiber/Protocol
From 2010.igem.org
(Difference between revisions)
Line 8: | Line 8: | ||
=== Grow up a culture of A.xylinus in media recommend by Open Wet Ware=== | === Grow up a culture of A.xylinus in media recommend by Open Wet Ware=== | ||
<html> | <html> | ||
- | <center><table><tr><td width="850" | + | <center><table><tr><td width="850" align="left"> |
<font size="3"><span style="text-decoration:underline">Material</font></span> | <font size="3"><span style="text-decoration:underline">Material</font></span> | ||
<ul><li>A.xylinus JCM strain 7664 | <ul><li>A.xylinus JCM strain 7664 | ||
Line 40: | Line 40: | ||
<li>The growth of A.xylinus is accompanied by the formation of a thick cellulose matrix within the media that must be removed before cells can be pelleted for a miniprep procedure. Simply vortex briefly to break up the cellulose into chunks and remove the cellulose chunks from the media with a pipette while carefully avoiding the removal of cells. | <li>The growth of A.xylinus is accompanied by the formation of a thick cellulose matrix within the media that must be removed before cells can be pelleted for a miniprep procedure. Simply vortex briefly to break up the cellulose into chunks and remove the cellulose chunks from the media with a pipette while carefully avoiding the removal of cells. | ||
<li>A.xylinus will grow well at room temperature in aerobic conditions.</ol> | <li>A.xylinus will grow well at room temperature in aerobic conditions.</ol> | ||
- | + | </td></tr></table></center> | |
Revision as of 08:57, 29 August 2010
![](https://static.igem.org/mediawiki/2010/9/9f/Re-main.png)
E.coli Fiber Project Protocol
Contents |
Grow up a culture of A.xylinus in media
Grow up a culture of A.xylinus in media recommend by Open Wet Ware
Material
Equipment
Procedure
Note
|
Grow up a culture of A.xylinus in media recommend by JCM
Material A.xylinus JCM strain 7664 1l of liquid Acetobacter media(JCM recommended) -Glucose - 100g -Yeast extract - 10g -CaCO3 30g -Distilled water - 1000 ml (If you are making plates, use the same protocol but add 15g of agar.) Equipment autoclave incubator scale bunsen burner flask(1l or500ml) plate spreader pipet pipet tip Procedure
Note
|
Grow up a culture of E.coli
Material Equipment Procedure |
Direct PCR
Material E.coli K12 colony 10µM/l forward Primer 5µl 10µM/l reverse Primer 5µl 10×EX taq buffer 10µl 2.5mM each dNTP mixture 10µl EX taq polymerase 1µl milli-Q 71µl Equipment thermal cycler vortex mixer PCR-tubes pipet pipet tip Procedure
|
Electrophoreses PCR productions
Material Equipment Procedure |
DNA purification from agarose gel with QIAGEN
Material QIAGEN(gel extraction kit) -QG buffer 300µl -PE buffer700µl -EB buffer 50µl -tube for column Equipment centrifuge heating plate pipet pipet tip Procedure 1. cut gel of electrophoreses 2. add pieces of gel to tubes 3. take 300µl QG buffer into tubes and dissolve at 50°C 4. add a solution of QG buffer and gel to tubes for column 5. centrifuge 15000rpm/1min 6. throw “flow-thru” away and take 700µl PE buffer 7. centrifuge 15000rpm/1min 8. throw flow-thru away 9. centrifuge 15000rpm/1min 10. change tube for column 11. add 50µl of EB buffer (aim to center of tube) 12. centrifuge 15000rpm/1min |