Paris/August 6

From 2007.igem.org

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==PCR on colonies==
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* P6 : again (1,7) + new (9,10,11,12)
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* P7 : again (1) + new (9,10,11,12,13,14,15)
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* P8 : again (1,3,5,7,8) + new (9,10,11)
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* P10 : again (old ones 1,5,6) + new (9,10,11,12,13)

Revision as of 18:27, 6 August 2007

Digestion reactions

Redigestion reactions were performed on several PCR products already digested. This was done because of owr persistant inability to clone any PCR product.

2x reaction mixes were prepared:

  • XbaI+PstI digestion reactions (buffer NEB3)
  • EcoRI+PstI (NEBU buffer)
  • SpeI+PstI (NEB2)
Digestion Products
Number Product Name Matrix Name Enzyme 1 Enzyme 2 Size Description vol. left for dig.
D19' BB dig. lox66DapAsubtilis lox66-DapAsubtilis P6' PCR product EcoRI PstI ~960 PCR product digested for cloning as a BioBrick 20
D21' bw-insert lox66-DapAsubtilis lox66-DapAsubtilis P6' PCR product XbaI PstI ~960 lox66-DapAsubtilis P6' PCR product 25
D27' lox71-ftsZ bw-insert lox71-ftsZ PCR product P7 XbaI PstI lox71-ftsZ(unmutated) ready for use as a backward insert 30
D28 RBS-DapAColi BB P8 (RBS-DapAColi) EcoRI PstI ~950 Biobrick 15
D29 RBS-DapAColi P8 (RBS-DapAColi) XbaI PstI ~950 Backward insert 13
D30 RBS-DapASubtilis BB P9 (RBS-DapASubtilis) EcoRI PstI ~950 Biobrick 14
D31 RBS-DapASubtilis P9 (RBS-DapASubtilis) XbaI PstI ~950 Backward insert 14
D32 Lox71-FtsA-FtsZ P10 Lox71-FtsA-FtsZ XbaI PstI ~2600 Backward insert - EcoRI site still presents : we can't use EcoRI 18
D34 lox66 L9' lox66 SpeI PstI Biobrick plasmid carrying lox66 digested as a BV 26
D35 lox71 L10' lox71 SpeI PstI Biobrick plasmid carrying lox71 digested as a BV 33
D36 lox71-KmR-lox66 P11 lox71-KmR-lox66 EcoRI PstI lox71-KmR-lox66 digested as a biobrick insert 28


Also, new digestion reactions were performed:

Digestion Products
Number Product Name Matrix Name Enzyme 1 Enzyme 2 Size Description
D18 BB dig. lox66DapAE.coli lox66-DapAE.coli P5 PCR product EcoRI PstI ~960 PCR product digested for cloning as a BioBrick
D20 bw-insert lox66-DapAE.coli lox66-DapAE.coli P5 PCR product XbaI PstI ~960 PCR product digested for usage as a backward insert
D20 bw-insert lox66-DapAE.coli lox66-DapAE.coli P5 PCR product XbaI PstI ~960 PCR product digested for usage as a backward insert
D20 bw-insert lox66-DapAE.coli lox66-DapAE.coli P5 PCR product XbaI PstI ~960 PCR product digested for usage as a backward insert
D20 bw-insert lox66-DapAE.coli lox66-DapAE.coli P5 PCR product XbaI PstI ~960 PCR product digested for usage as a backward insert
D20 bw-insert lox66-DapAE.coli lox66-DapAE.coli P5 PCR product XbaI PstI ~960 PCR product digested for usage as a backward insert
D20 bw-insert lox66-DapAE.coli lox66-DapAE.coli P5 PCR product XbaI PstI ~960 PCR product digested for usage as a backward insert
D20 bw-insert lox66-DapAE.coli lox66-DapAE.coli P5 PCR product XbaI PstI ~960 PCR product digested for usage as a backward insert

PCR on colonies

  • P6 : again (1,7) + new (9,10,11,12)
  • P7 : again (1) + new (9,10,11,12,13,14,15)
  • P8 : again (1,3,5,7,8) + new (9,10,11)
  • P10 : again (old ones 1,5,6) + new (9,10,11,12,13)