Organismal Fitness and Auxotroph Complementation Selection

Creative Biostructure can provide custom screening and selection services based on organismal fitness and auxotroph complementation methods for protein evolution and engineering assays as well as other applications.

The directed screening and selection of mutant library is one of the most critical steps in protein evolution and engineering process. Genetic screenings and functional protein selections can be achieved by multiple methods. Organismal fitness and survival is one of the most popular basic methods for directed evolution. Active library members can enable organisms that containing their corresponding genes to survive and replicate under a certain directed selection environment. For example, antibiotic resistance is the most straightforward activity applied for the directed selection assays of E. coli. Enzymes that neutralize or export antibiotics have evolved and be selected in various studies. By linking the desired activity to the expression of an antibiotic resistance gene, we can also evolve other proteins with the antibiotic survival selections and screenings.

Auxotroph complementation can also form the basic selection and screening methods, especially for proteins or enzymes that are involved in the metabolism. An auxotroph is an organism that can't synthesize a particular organic compound required for its growth. The auxotroph complementation selection methods take the auxotroph complementation as an ideal selectable marker for directed protein evolution selection and screening. For example, the metabolism of Xylose has been applied in protein evolution assays as an important target because of its limitation effects on the conversion of lignocellulose biomass into ethanol for use in biofuels. Media containing xylose as the sole carbon source is used for selection. Organisms that contain genes encoding enzymes that better utilize this energy source are enhanced in growth and selected out.

As a basic selection and screening method, organismal fitness and auxotroph complementation based method is relatively simple in concept. The throughout can rise up to a level of 10⁸ or 10¹⁰ (Depends on different hosts). Selectable markers (e.g. antibiotic resistance) metabolic genes and suppressors of toxic genes are ideal targets for organismal fitness and auxotroph complementation based strategies.

With state-of-art platforms and years of experience in the field of protein evolution and engineering, Creative Biostructure can provide our customers the custom organismal fitness and auxotroph complementation based screening and selection services as well as other methods. Our science team will develop the optimal strategy for each individual case with organismal fitness and auxotroph complementation based methods alone or combined with other methods.  

Creative Biostructure also provides other selection and screening services based on various different methods. Welcome to contact us for more details.

Reference:
Borsuk S, Mendum T A, Fagundes M Q, et al. Auxotrophic complementation as a selectable marker for stable expression of foreign antigens in Mycobacterium bovis BCG[J]. Tuberculosis, 2007, 87(6): 474-480.
Packer M S, Liu D R. Methods for the directed evolution of proteins[J]. Nature Reviews Genetics, 2015, 16(7): 379-394.