Difference between revisions of "Plasmid backbones/Version 5/Features"
 
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#A complete BioBrick® cloning site for easy cloning and assembly of BioBrick® parts.
 
#A complete BioBrick® cloning site for easy cloning and assembly of BioBrick® parts.
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#A ''ccdB'' positive selection marker within the BioBrick® cloning site.  The ''ccdB'' gene ensures that when assembling two BioBrick® parts together, the uncut plasmid vector is not transformed.  However, inclusion of the ''ccdB'' gene means that these vectors must be propagated in a ''ccdB'' tolerant strain, such as ''E. coli'' strain DB3.1 ([[Part:BBa_V1005|BBa_V1005]]).
 
 
#A low or medium copy replication origin to reduce consumption of cellular resources during device or system operation.
 
#A low or medium copy replication origin to reduce consumption of cellular resources during device or system operation.
 
#Forward and reverse terminators both upstream and downstream of the BioBrick® cloning site to insulate the vector from read-through transcription originating in the cloned BioBrick® device or system and to insulate the cloned BioBrick® system from the vector.
 
#Forward and reverse terminators both upstream and downstream of the BioBrick® cloning site to insulate the vector from read-through transcription originating in the cloned BioBrick® device or system and to insulate the cloned BioBrick® system from the vector.
 
#Stop codons in all reading frames to prevent inadvertent translation into or out of the BioBrick cloning site.
 
#Stop codons in all reading frames to prevent inadvertent translation into or out of the BioBrick cloning site.
 
#Primer binding sites for the standard BioBrick® verification primers VF2 ([[Part:BBa_G00100|BBa_G00100]]) and VR ([[Part:BBa_G00101|BBa_G00101]]).  These primers are located for convenient sequencing and [http://openwetware.org/wiki/Colony_PCR screening by colony PCR] of cloned BioBrick® devices and systems.
 
#Primer binding sites for the standard BioBrick® verification primers VF2 ([[Part:BBa_G00100|BBa_G00100]]) and VR ([[Part:BBa_G00101|BBa_G00101]]).  These primers are located for convenient sequencing and [http://openwetware.org/wiki/Colony_PCR screening by colony PCR] of cloned BioBrick® devices and systems.
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Plasmid backbones are distributed by the Registry with a default insert.  There are just a handful of default plasmid inserts used in the Registry.  All of the available plasmid backbones include the RFP expression cassette ([[Part:BBa_J04450|BBa_J04450]]) as the default plasmid insert within the BioBrick® cloning site.  The RFP cassette enables red/white screening of colonies and ensures that when assembling two BioBrick® parts together, the uncut plasmid (or a plasmid that get the RFP cassette as insert) will be red on the plate whereas the correct colonies will be white.
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Plasmid backbones with the plasmid insert of [[Part:BBa_I52002|BBa_I52002]] also include a high copy replication origin in the default insert.  Thus, these plasmid backbones are easily [http://openwetware.org/wiki/Miniprep/Qiagen_kit purified in large quantities].  Cloning or assembly of a BioBrick® part into the BioBrick® cloning site of the plasmid backbone eliminates the default insert returning the plasmid to the control of the replication origin in the plasmid backbone.

Latest revision as of 15:41, 23 September 2012

  1. A complete BioBrick® cloning site for easy cloning and assembly of BioBrick® parts.
  2. A low or medium copy replication origin to reduce consumption of cellular resources during device or system operation.
  3. Forward and reverse terminators both upstream and downstream of the BioBrick® cloning site to insulate the vector from read-through transcription originating in the cloned BioBrick® device or system and to insulate the cloned BioBrick® system from the vector.
  4. Stop codons in all reading frames to prevent inadvertent translation into or out of the BioBrick cloning site.
  5. Primer binding sites for the standard BioBrick® verification primers VF2 (BBa_G00100) and VR (BBa_G00101). These primers are located for convenient sequencing and [http://openwetware.org/wiki/Colony_PCR screening by colony PCR] of cloned BioBrick® devices and systems.

Plasmid backbones are distributed by the Registry with a default insert. There are just a handful of default plasmid inserts used in the Registry. All of the available plasmid backbones include the RFP expression cassette (BBa_J04450) as the default plasmid insert within the BioBrick® cloning site. The RFP cassette enables red/white screening of colonies and ensures that when assembling two BioBrick® parts together, the uncut plasmid (or a plasmid that get the RFP cassette as insert) will be red on the plate whereas the correct colonies will be white.

Plasmid backbones with the plasmid insert of BBa_I52002 also include a high copy replication origin in the default insert. Thus, these plasmid backbones are easily [http://openwetware.org/wiki/Miniprep/Qiagen_kit purified in large quantities]. Cloning or assembly of a BioBrick® part into the BioBrick® cloning site of the plasmid backbone eliminates the default insert returning the plasmid to the control of the replication origin in the plasmid backbone.