Part:BBa_K3945014

Xpress: T7-Compatible Expression Vector for E. coli with A GST solubility Tag

Usage and Biology

Xpress is an easy-to-use expression vector for E. coli BL21 system designed by the Schryvers’s lab. With a T7 promoter, this vector is easily inducible with IPTG or lactose. Two unique features set this expression vector apart from the other E. coli vectors: a quick screening system allows for fast cloning of genetic constructs into E. coli and a Glutathione S-transferase tag (GST) for easy expression of difficult proteins.

Design

Quick Screening: Screening for positive colonies following cloning is often a long and tedious task that often requires screening of many colonies. With the help of a unique feature on the Xpress vector, screening for a positive colony could be done in one single step. Xpress contains two selection markers: a kanamycin resistance gene and an ampicillin resistance gene. However, upon cloning of the insert into the appropriate spot in the multiple cloning sites, the kanamycin resistance gene will be knocked out. As such, successfully cloned colonies will only be resistant to ampicillin whereas colonies that have not been engineered properly will retain their kanamycin resistance. By streaking colonies on ampicillin and kanamycin plates, successful and unsuccessful colonies could be easily differentiated from one another as the successful colonies are expected to only grow on the ampicillin plates while all other negative colonies will show growth on both kanamycin and ampicillin plate. Thus, in this one simple step Xpress allows for fast screening of positive colonies.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 33
  Illegal XbaI site found at 1908
  Illegal PstI site found at 2955
  Illegal PstI site found at 3842
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 33
  Illegal PstI site found at 2955
  Illegal PstI site found at 3842
  Illegal NotI site found at 3
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 33
  Illegal BamHI site found at 757
  Illegal XhoI site found at 912
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 33
  Illegal XbaI site found at 1908
  Illegal PstI site found at 2955
  Illegal PstI site found at 3842
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 33
  Illegal XbaI site found at 1908
  Illegal PstI site found at 2955
  Illegal PstI site found at 3842
  Illegal NgoMIV site found at 2201
  Illegal NgoMIV site found at 2598
  Illegal NgoMIV site found at 2608
  Illegal NgoMIV site found at 2734
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal BsaI site found at 3694
  Illegal SapI.rc site found at 160
  Illegal SapI.rc site found at 2705