A modified PCR system for amplifying beta-ketoacyl-ACP synthase gene fragments with DNA from Streptomyces luteogriseus.
Streptomyces luteogriseus strain 099, producing a new type of macrolide antibiotic with anti-coxB6 virus and anti-HIV protease activities, was isolated from soil. PCR was optimized to amplify beta-ketoacyl-ACP synthase (KS) genes. The system was optimized around the use of higher concentrations of DMSO (15% vs. 10% v/v) and dNTP (500 microM vs. 50-200 microM) and a lower annealing temperature (55 degrees C vs. 60-70 degrees C) than the normal PCR method used to amplify high GC content DNA.
Yu
Feng-Ming
FM
Department of Pharmaceutical Engineering, Tianjin University, P.O. Box 6888, 300072, Tianjin, PR China.
Jiang
Xin
X
Wu
Jin-Chuan
JC
Yuan
Ying-Jin
YJ
D015099:3-Oxoacyl-(Acyl-Carrier-Protein) Synthase / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics*; D017320:HIV Protease Inhibitors / Q000378:metabolism; D018942:Macrolides / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA; D013302:Streptomyces / Q000201:enzymology / Q000235:genetics*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D017320:HIV Protease Inhibitors; D018942:Macrolides; D015099:3-Oxoacyl-(Acyl-Carrier-Protein) Synthase