Parallel capillary electrophoresis for the quantitative screening of fermentation broths containing natural products.
Directed molecular evolution is a recursive process of controlled genetic diversification and functional screening. The success of this approach is dependent on both the quality of the genetic diversity and the ability to accurately screen a large population of individual genetic variants for those having improved function. In this paper, the application of parallel capillary electrophoresis to rapidly quantitate lovastatin production levels by Aspergillus terreus mutants is described. A parallel 96 capillary instrument analyzed 900 samples in 8 h. with a 100 mM MES at pH 5.2 running buffer. In this manner, the fermentation broths of thousands of mutated strains were efficiently and inexpensively screened for increased lovastatin production. The ability to develop high-throughput methods to both separate and quantitate the components of complex mixtures greatly facilitates the ability to apply evolutionary engineering methods to complex biological systems.
Kittell
Justin
J
Codexis Inc., 200 Penobscott Drive, Redwood City, CA 94063, USA.
Borup
Birthe
B
Voladari
Rama
R
Zahn
Ken
K
D001230:Aspergillus / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D045424:Complex Mixtures / Q000032:analysis*; D003470:Culture Media / Q000032:analysis* / Q000378:metabolism; D019075:Electrophoresis, Capillary / Q000379:methods*; D005285:Fermentation / Q000502:physiology; D024861:Genetic Enhancement / Q000379:methods; D008148:Lovastatin / Q000032:analysis* / Q000378:metabolism*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D018511:Systems Integration
D045424:Complex Mixtures; D003470:Culture Media; D008148:Lovastatin