Difference between revisions of "Part:BBa K1223012"

 
 
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<partinfo>BBa_K1223012 short</partinfo>
 
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===Design Notes===
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just need to assemble with selection gene of choice to get any plasmid from the pKD family - pKD46 carries the bla gene (ampicillin resistance), pKD78 carries the cat gene(chloramphenicol resistance), pKD119 carries the tet gene (tetracycline resistance)or create new plasmid with kanamicin resistance.
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temperature sensitive origin of replication. grow the bacteria at 30C to keep and amplify the plasmid or grow at 42C to eliminate it.
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===Source===
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genbank.
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===References===
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Kirill A. Datsenko and Barry L. Wanner
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One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products
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PNAS 2000 97 (12) 6640-6645; published ahead of print May 30, 2000, doi:10.1073/pnas.120163297
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===Usage and Biology===
 
===Usage and Biology===
  

Latest revision as of 13:51, 12 October 2013

pKD46 functional unit

pKD46 functional part - the plasmid backbone without antibiotic resistance.

pKD plasmids are carrying the Lambda Red recombineering system. this system is highly efficient in making indels in E.coli genome.

just need to assemble with selection gene of choice to get any plasmid from the pKD family - pKD46 carries the bla gene (ampicillin resistance), pKD78 carries the cat gene(chloramphenicol resistance), pKD119 carries the tet gene (tetracycline resistance)or create new plasmid with kanamicin resistance.

the part includes: - AraC repressor CDS. - beta gamma and exo genes of the lambda bacteriophage regulated by AraBAD promoter (arabinose induction). - OriR101 replication origin - temperature sensitive origin. grow the bacteria at 30C to keep and amplify the plasmid or grow at 42C to eliminate it.

Kirill A. Datsenko and Barry L. Wanner One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products PNAS 2000 97 (12) 6640-6645; published ahead of print May 30, 2000, doi:10.1073/pnas.120163297


Design Notes

just need to assemble with selection gene of choice to get any plasmid from the pKD family - pKD46 carries the bla gene (ampicillin resistance), pKD78 carries the cat gene(chloramphenicol resistance), pKD119 carries the tet gene (tetracycline resistance)or create new plasmid with kanamicin resistance.

temperature sensitive origin of replication. grow the bacteria at 30C to keep and amplify the plasmid or grow at 42C to eliminate it.


Source

genbank.

References

Kirill A. Datsenko and Barry L. Wanner One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products PNAS 2000 97 (12) 6640-6645; published ahead of print May 30, 2000, doi:10.1073/pnas.120163297


Usage and Biology

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 1207
    Illegal EcoRI site found at 2716
    Illegal SpeI site found at 4505
    Illegal PstI site found at 2211
    Illegal PstI site found at 2458
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 1207
    Illegal EcoRI site found at 2716
    Illegal SpeI site found at 4505
    Illegal PstI site found at 2211
    Illegal PstI site found at 2458
    Illegal NotI site found at 4824
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 1207
    Illegal EcoRI site found at 2716
    Illegal BamHI site found at 1146
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 1207
    Illegal EcoRI site found at 2716
    Illegal SpeI site found at 4505
    Illegal PstI site found at 2211
    Illegal PstI site found at 2458
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 1207
    Illegal EcoRI site found at 2716
    Illegal SpeI site found at 4505
    Illegal PstI site found at 2211
    Illegal PstI site found at 2458
    Illegal AgeI site found at 981
    Illegal AgeI site found at 2638
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 963