Chimeragenesis

Coco, W.M. (2003) RACHITT gene family shuffling by random chimeragenesis on transient templates. Methods in Molecular Biology (Clifton, NJ), 231, 111-127. 

Volkov, A.A., Shao, Z. and Arnold, F.H. (1999) Recombination and chimeragenesis by in vitro heteroduplex formation and in vivo repair. Nucleic Acids Research, 27, el8. 

Coco, W. M., RACHITT Gene family shuffling by Random Chimeragenesis on Transient Templates. Methods. Mol.Biol., 2003. 231 pp. 111-127. 

Protein engineering chimeragenesis and site-directed mutagenesis... 

There are two basic considerations when attempting SDM. One is to determine the amino acids that should be mutated and the other is to decide what to replace them with. The first question is, of course, dependant upon information gathered from previous experimentation in order to target residues that are appropriate. Such information may be derived from biochemical techniques. For instance, in our binding site studies, we have specifically mutated amino acids that had previously shown to be covalently labeled by photoactive ligands. Additionally, we have used comparisons between the sequences of different receptor subunits that correlate with receptor function to identify domains of interest. Chimeragenesis, the technique described in the first half of this chapter, can provide important information in this regard. Obviously, those proteins for which a detailed structural model is available will lend themselves to more rational substitutions. 

Chimeragenesis and SDM are powerful techniques that can be used to investigate the complex relationships between protein structure and function. The methods detailed here are relatively simple to perform and can be carried out in a short period of time. They are applicable to any protein type for which the cDNA is available and can be modified for many different purposes in protein engineering. 

Brock BJ, Waterman MR. 2000. The use of random chimeragenesis to study structure/function properties of rat and human P450cl7. Arch Biochem Biophys 373(2) 401 Colquhoun D. 1998. Binding, gating, affinity and efficacy the interpretation of structure-activity relationships for agonists and of the effects of mutating receptors. Br J Pharmacol 125(5) 924. 

Protein Engineering Chimeragenesis and Site-Directed Mutagenesis. 423... 

Random Chimeragenesis on Transient Templates (RAC HITT) all generate library members are shuffled more crossovers than possible with PCR methods Coco, 2001... 

Figure 11.6 Random Chimeragenesis on Transient Templates (RACHITT) with mutagenesis (Gibbs, private communication).

SISDC Sequence-independent site-directed chimeragenesis... 

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