Difference between revisions of "Featured Parts:Conjugation parts"

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===Experimental and Technical===
 
===Experimental and Technical===
 
* Conjugation is most often carried out in the lab by either of the two following processes
 
* Conjugation is most often carried out in the lab by either of the two following processes
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##Liquid Media: Recipient culture and donor culture are grown separately  in liquid media overnight at 37C.  Add both cultures to the same sterile container.  Allow a half hour to two hours for conjugation to occur.  Do not disturb (ie. shake) the container--it will cause the pili to shear and thus prevent conjugation.  (Note: this method is suitable for the F-type conjugal plasmid carriers, but not the R-type conjugal plasmid, whose pili are not as robust)
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#Liquid Media: Recipient culture and donor culture are grown separately  in liquid media overnight at 37C.  Add both cultures to the same sterile container.  Allow a half hour to two hours for conjugation to occur.  Do not disturb (ie. shake) the container--it will cause the pili to shear and thus prevent conjugation.  (Note: this method is suitable for the F-type conjugal plasmid carriers, but not the R-type conjugal plasmid, whose pili are not as robust)
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##Solid Media: Recipient culture and donor cultures are grown separately overnight on solid agar plates.  Mix cells together on the same sterile agar plate using an innoculating loop, streaking, etc.  This method is suitable for all types of conjugative plasmids (F, R, etc).
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#Solid Media: Recipient culture and donor cultures are grown separately overnight on solid agar plates.  Mix cells together on the same sterile agar plate using an innoculating loop, streaking, etc.  This method is suitable for all types of conjugative plasmids (F, R, etc).
  
 
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Revision as of 07:14, 17 April 2006

The Parts

BBa_J01002 "OriTf" - Origin of Transfer for the F-type conjugative plasmid
BBa_J01003 "OriTr" - Origin of Transfer for the R-type conjugative plasmid
BBa_J01000 "TraJf" - Initiates expression of mating characteristic in the F-type conjugative plasmid
BBa_J01001 "TraJr" - Initiates mating characteristics in the R-type conjugative plasmid

Background/About Conjugation

conjugating cells connected by a pilus

Bacterial conjugation is the transfer of genetic material between cells--a process often defined as the bacterial equivalent of mating. This behaviour is characterized by the presence of a specialized plasmid (small, circular piece of transferrable DNA most often found within bacteria) known as a "conjugative plasmid" .
The conjugative plasmid holds a distinct and specialized set of coding regions including the following

  • OriT (Origin of Transfer): Unlike other plasmids, the conjugative plasmid has its own origin of transfer.
  • TraJ (Transfer genes): The activation of this gene sets off a cascade of other plasmid genes (and thus corresponding protein expressions) which act in concert to form "mating" characteristics in the host bacterial cell. Such salient characteristics include:
    • the growth of a pilus (a long whip-like apparatus used for genetic transfer between cells)
    • formation of "surface exclusion proteins" which prevents the cell containing the conjugative plasmid from mating with other conjugative plasmids of its type

Two of the most well-studied and characterized types of conjugative plasmids are types "F" and "R". The employed nomenclature for conjugative plasmid genes is the gene name first with the type afterwards. For example, the "TraJr" or "TraJR" gene denotes the "TraJ" gene of the "R" type conjugative plasmid.

Experimental and Technical

  • Conjugation is most often carried out in the lab by either of the two following processes
  1. Liquid Media: Recipient culture and donor culture are grown separately in liquid media overnight at 37C. Add both cultures to the same sterile container. Allow a half hour to two hours for conjugation to occur. Do not disturb (ie. shake) the container--it will cause the pili to shear and thus prevent conjugation. (Note: this method is suitable for the F-type conjugal plasmid carriers, but not the R-type conjugal plasmid, whose pili are not as robust)
  2. Solid Media: Recipient culture and donor cultures are grown separately overnight on solid agar plates. Mix cells together on the same sterile agar plate using an innoculating loop, streaking, etc. This method is suitable for all types of conjugative plasmids (F, R, etc).

More Information/References

  1. Frost, Ipp en-Ihler, Skurray. Microbial Rev 1994. Analysis and Sequence of the F Plasmid [http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&dopt=Abstract&list_uids=7915817&query_hl=2&itool=pubmed_docsum Pubmed]
  2. Wikipedia. "Bacterial Conjugation" [http://en.wikipedia.org/wiki/Bacterial_conjugation Link]
  3. Berkeley iGEM 2005 [http://parts.mit.edu/wiki/index.php/Berkeley_2005 Project]