Difference between revisions of "Part:BBa K1223001:Design"

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===Design Notes===
 
===Design Notes===
 
This part was designed to fulfill the self destruct system of P.A.S.E 1. It contains a toxin system based on phage lysis system of holin (BBa_K112805) and lysozyme (BBa_K112301). Holin protein causes "pores" in the inner membrane, which allows lysozyme to access and break down the peptidoglycan of the cell wall, causing lysis and eventually death. The toxins are regulated by cI regulated promoter (BBa_R0051). This part is designed to integrate into the cell's genome via homologous recombination and therefore it contains homologous regions at its ends. Kanamycin resistance was added for selectivity. Therefore, when transforming in bacteria only the cells that have gone through double recombination with the insert will survive.   
 
This part was designed to fulfill the self destruct system of P.A.S.E 1. It contains a toxin system based on phage lysis system of holin (BBa_K112805) and lysozyme (BBa_K112301). Holin protein causes "pores" in the inner membrane, which allows lysozyme to access and break down the peptidoglycan of the cell wall, causing lysis and eventually death. The toxins are regulated by cI regulated promoter (BBa_R0051). This part is designed to integrate into the cell's genome via homologous recombination and therefore it contains homologous regions at its ends. Kanamycin resistance was added for selectivity. Therefore, when transforming in bacteria only the cells that have gone through double recombination with the insert will survive.   
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[[File:Example.jpg]]
 
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The part was characterized through sequencing and restriction digest with BamHI and EcorRV.
 
The part was characterized through sequencing and restriction digest with BamHI and EcorRV.
 +
 +
[[File:Example2.jpg]]
  
 
lane1: undigested pUC57-P.A.S.E.1 cassette
 
lane1: undigested pUC57-P.A.S.E.1 cassette
 +
 
lane2: pUC57-P.A.S.E.1 cassette digested with BamHI and EcorRV.
 
lane2: pUC57-P.A.S.E.1 cassette digested with BamHI and EcorRV.
−
lane3: DS5000 Ladder
 
  
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[[File:Example2.jpg]]
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lane3: DS5000 Ladder
  
 
===Source===
 
===Source===

Revision as of 15:03, 1 October 2013

P.A.S.E. 1 cassette


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 13
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

This part was designed to fulfill the self destruct system of P.A.S.E 1. It contains a toxin system based on phage lysis system of holin (BBa_K112805) and lysozyme (BBa_K112301). Holin protein causes "pores" in the inner membrane, which allows lysozyme to access and break down the peptidoglycan of the cell wall, causing lysis and eventually death. The toxins are regulated by cI regulated promoter (BBa_R0051). This part is designed to integrate into the cell's genome via homologous recombination and therefore it contains homologous regions at its ends. Kanamycin resistance was added for selectivity. Therefore, when transforming in bacteria only the cells that have gone through double recombination with the insert will survive.

The part was characterized through sequencing and restriction digest with BamHI and EcorRV.

Example2.jpg

lane1: undigested pUC57-P.A.S.E.1 cassette

lane2: pUC57-P.A.S.E.1 cassette digested with BamHI and EcorRV.

lane3: DS5000 Ladder

Source

part is synthetic.

holin and lysozime originates from lambda bacteriophage.

the homology regions were taken from E.coli BL21 genome

References