Vector adaptation to the Staby(r) technology

The GetStabyTM application allows easy addition of Delphi-Genetics' stabilization technology into your favourite vector. The techology is compatible with any expression system. Using this technology, your vectors are perfectly stabilized even without antibiotics.

Principle

Some inserts, when cloned into a classical vector, may cause instability of this vector resulting in low yield of DNA or protein production (for protein expression vectors). The reasons for this instability are, for example, the toxicity of the insert and/or the secondary structure and/or the size of the vector. The GetStabyTM application provides, on one hand, a plasmid containing an antidote gene (ccdA) under the control of a constitutive promoter and, on the other hand, an E. coli strain (CYS22) that contains, in its chromosome, the selection gene (ccdB) under the control of a promoter strongly repressed in the presence of the plasmid. Contrary to classical strains, if some CYS22 bacteria lose the vector, they will not obtain a selective (growth speed) advantage, but will die. In practice, this means that at any time during bacterial growth, 100% of the CYS22 bacteria will carry the vector even in the absence of antibiotics. These bacteria will yield high amounts of DNA or protein. Adding the StabyTM cassette (the DNA fragment encoding the CcdA antidote protein) to a vector results in stabilization of this vector in CYS22 (or any other strain containing the ccdB-selection gene in its chromosome).

The StabyTM cassette is isolated from the pGetStabyA vector using restriction sites or PCR amplification.

The insertion of the StabyTM cassette into any vector is easily selected using the antidote properties of this cassette and the CYS22 bacteria (encoding the CcdB selection protein). Then, the stabilization cassette linked to any part of the vector can be easily exported to other vectors and selected using the same properties.

Service

We provide a GetStabyTM services that allows you to stabilize your own vectors. It is compatible with any expression system. Contact us for more information

Benefits of the GetStaby application

  • The DNA or protein production is increased using the stabilized vector (3 to 5 times);
  • Plasmids are perfectly stabilized without the use of antibiotics;
  • The insertion of the stabilization cassette into any vector is easily selected using the antidote properties;
  • When performing protein expression, the background of “parasite proteins” is reduced;
  • The GetStabyTM system is compatible with any vector;
  • The GetStabyTM system is usable in any culture medium (LB, SOC, SOB, YT or minimal media supplemented ith carbon source such as glucose or
    glycerol,...);
  • When using AmpR plasmids, no “satellite” colonies are observed even after extended incubation;
  • The stabilization cassette (linked to any part of the vector) is easily re-exported to other vectors.