Difference between revisions of "Part:BBa J70027:Design"

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(→‎Design Notes)
 
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The start of J70027 matches the origin start in the pSci1 plasmid.
 
The start of J70027 matches the origin start in the pSci1 plasmid.
 +
 +
There appears to be a frame shift mutation at base 659, shifting the PE gene.  This is likely fatal to replication.
 +
Re-PCR from the original S. citri genomic DNA sample and re-transform.
  
 
===Source===
 
===Source===

Latest revision as of 19:20, 17 November 2010

Mesoplasma florum plasmid origin


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 372


Design Notes

No native restriction sites. Added biobrick ends.

The gene for PE contains no CGG codons.

The primer names are for J70023 -- this is a shortened version

GTTTCTTCGAATTCGCGGCCGCTTCTAGAGtaattataaaaacaccttattttac,J00023-F2

GTTTCTTCCTGCAGCGGCCGCTACTAGTAttgggagtgcggggtttggggtgc,J00023-R

The start of J70027 matches the origin start in the pSci1 plasmid.

There appears to be a frame shift mutation at base 659, shifting the PE gene. This is likely fatal to replication. Re-PCR from the original S. citri genomic DNA sample and re-transform.

Source

Genbank AJ969070

PCR from S. citri genomic DNA, kind gift of J. Renaudin, using J70023-F2 and J70023-R primers.

References

Breton M, Duret S, Arricau-Bouvery N, Béven L, Renaudin J. Characterizing the replication and stability regions of Spiroplasma citri plasmids identifies a novel replication protein and expands the genetic toolbox for plant-pathogenic spiroplasmas. Microbiology. 2008 Oct;154(Pt 10):3232-44. PMID: 18832328

Saillard C, Carle P, Duret-Nurbel S, Henri R, Killiny N, Carrère S, Gouzy J, Bové JM, Renaudin J, Foissac X. The abundant extrachromosomal DNA content of the Spiroplasma citri GII3-3X genome. BMC Genomics. 2008 Apr 28;9:195. PMID: 18442384

Berho N, Duret S, Danet JL, Renaudin J. Plasmid pSci6 from Spiroplasma citri GII-3 confers insect transmissibility to the non-transmissible strain S. citri 44. Microbiology. 2006 Sep;152(Pt 9):2703-16. PMID: 16946265