Part:pSB3C01

pEven1 Loop Vector based on pSB3C5

pEven1 Loop Vector based on pSB3C5

iGEM Type IIS Vectors

pEven Loop Vectors (for Level 2)

When four Level 1 parts (TUs) are assembled into a Multi-Transcriptional Unit (MTU) into the following plasmid backbones (with BBa_J04455), the MTU will be flanked by BsaI restriction sites and these 4 bp Fusion Sites.

Registry Name	Loop Name	Fusion Site 5'	MTU	Fusion Site 3'
pSB3C01	pEven1	GGAG	MTU 1	TACT
pSB3C02	pEven2	TACT	MTU 2	AATG
pSB3C03	pEven3	AATG	MTU 3	GCTT
pSB3C04	pEven4	GCTT	MTU 4	CGCT

Note:

• The documented sequence shows an additional BsaI site. This would impact assembly out of (but not into) this set of vectors. A new set of vectors should be created or used in the long term.
• Currently, Registry samples of these plasmid backbones have not been fully sequenced. Only their prefix and suffix along with their insert BBa_J04455 have been verified.

The iGEM Type IIS assembly standard is based on MoClo and Loop

• A Modular Cloning System for Standardized Assembly of Multigene Constructs
• Loop assembly: a simple and open system for recursive fabrication of DNA circuits

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2713
  Illegal PstI site found at 12
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2713
  Illegal NheI site found at 1399
  Illegal PstI site found at 12
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2713
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2713
  Illegal PstI site found at 12
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2713
  Illegal PstI site found at 12
  Illegal AgeI site found at 990
  Illegal AgeI site found at 1313
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal BsaI site found at 285
  Illegal BsaI site found at 2719
  Illegal BsaI.rc site found at 6
  

Uppsala 2020's Improvement

The illegal BsaI site was removed by PCR mutagenesis to obtain the new pEven1 plasmid pSB3C11 (BBa_K3425001). This new plasmid was used for various clonings and it works as expected.