Template:Team:KULeuven/3 September 2009/VanillinProduction

From 2009.igem.org

(Difference between revisions)
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===NOT PCR===
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* electroporation of SAMS+TER ligation (from tuesday ligation... the one that looked bad....)
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* Miniprepped the 4 colonies from SAMS
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| border ="1" align="center"
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!| part || concentration || 260/280 || 260/230
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|- align="center"
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| SAMS 1 || 35,4 || 1,97 || 1,18
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|- align="center"
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| SAMS 2 || 49,3 || 2,00 || 0,97
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|- align="center"
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| SAMS 3 || 62,0 || 1,97 || 2,65
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|- align="center"
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| SAMS 4 || 78,3 || 1,85 || 1,43
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|- align="center"
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| Terminator || 181,4 || ||
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|-
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|}
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* Restriction digest of SAMS
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* Gel electrophorese of restriction from ech, fcs, sam5 and sam8
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** Results looks great (woohoo!)
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** Cut and purified from gel
 +
| border ="1" align="center"
 +
!| gene || concentration || 260/280 || 260/230
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|- align="center"
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| Sam5 || 8,2 || 1,62 || 0,02
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|- align="center"
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| Sam8 || 1,4 || 3,04 || 0,01
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|- align="center"
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| ech || 10,1 || 1,50 || 0,05
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|- align="center"
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| fcs || 5,2 || 1,61 || 0,03
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|-
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|}
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*Used ech and fcs to ligate using 5 μl from ech and 23,5 μl from fcs. There was not enough Sam8 for ligation. Redo restriction at some point in the future...
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 +
==== PCR ====
==== PCR ====
* The PCR from yesterday went very well, we had a huge amount of amplified biobrick DNA of the different parts. Agarose gel electrophoresis was used to test if the different pieces of DNA created by PCR had the correct length.
* The PCR from yesterday went very well, we had a huge amount of amplified biobrick DNA of the different parts. Agarose gel electrophoresis was used to test if the different pieces of DNA created by PCR had the correct length.

Revision as of 15:00, 4 September 2009

NOT PCR

  • electroporation of SAMS+TER ligation (from tuesday ligation... the one that looked bad....)
  • Miniprepped the 4 colonies from SAMS

| border ="1" align="center" !| part || concentration || 260/280 || 260/230 |- align="center" | SAMS 1 || 35,4 || 1,97 || 1,18 |- align="center" | SAMS 2 || 49,3 || 2,00 || 0,97 |- align="center" | SAMS 3 || 62,0 || 1,97 || 2,65 |- align="center" | SAMS 4 || 78,3 || 1,85 || 1,43 |- align="center" | Terminator || 181,4 || || |- |}

  • Restriction digest of SAMS
  • Gel electrophorese of restriction from ech, fcs, sam5 and sam8
    • Results looks great (woohoo!)
    • Cut and purified from gel

| border ="1" align="center" !| gene || concentration || 260/280 || 260/230 |- align="center" | Sam5 || 8,2 || 1,62 || 0,02 |- align="center" | Sam8 || 1,4 || 3,04 || 0,01 |- align="center" | ech || 10,1 || 1,50 || 0,05 |- align="center" | fcs || 5,2 || 1,61 || 0,03 |- |}

  • Used ech and fcs to ligate using 5 μl from ech and 23,5 μl from fcs. There was not enough Sam8 for ligation. Redo restriction at some point in the future...


PCR

  • The PCR from yesterday went very well, we had a huge amount of amplified biobrick DNA of the different parts. Agarose gel electrophoresis was used to test if the different pieces of DNA created by PCR had the correct length.
gene concentration 260/280 260/230
Sam5 361,6 1,91 2,13
Sam8 323,8 1,92 2,06
ech 236,5 1,91 2,24
fcs 321,4 1,90 2,22
  • Amplified DNA was purified and cut with different restriction enzymes. After restriction, DNA was purified again before ligation.
gene concentration 260/280 260/230
Sam5 21,8 1,65 1,33
Sam8 17,1 1,56 1,16
ech 13,8 1,78 1,46
fcs 20,9 1,70 0,20
  • After the restriction, Sam5, Sam8 and terminator were ligated in a three-way ligation. The same was done for ech, fcs and terminator. We also started a ligation of just Sam5 and Sam8 and ech and fcs.
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