Multiple-resistance Screening On Insecticidal Microbial Strain

The novel spinosyns are very similar to spinosyns, differing in the length of the side chain at C-21. The distinctive feature was the presence of the typical ethyl group at the C-21position of the macrocyclic ring system. To date, more than30factors have been isolated and characterized with the primary factors21-novel spinosyn A and D. Compared with spinosad, pogonin have a broader insecticidal spectrum and higher insecticidal activity, further study on it will become very important for research of biological insecticides. The performance of novel spinosyns is better than spinosad, but the biggest problem is the low yield, a long way for extensive industrialization produce that is hoped for.Drug-resistance mutation of microorganisms reflects structure and function changes of the ribosome and RNA polymerase. These changes can have significant effects on secondary metabolism in the mutant strain. Ribosome engineering mainly obtain the specific resistance mutations through the resistance screening method, related site of ribosomal or RNA polymerase structural changes caused by mutation not only affects the protein synthesis, but also significantly impact the synthesis of secondary metabolites. Though screening drug-resistance label can obtain the forward mutants efficiently, the general expression for the increase in production of secondary metabolites, or biosynthesis capacity, metabolic rate, stability of inheritance are significantly improved and enhanced.Based on the above, here we first determined the growth curve and the fermentation process curves of S. pogona, and then obtained a dual-resistant mutant SG58by ribosome engineering technique, the relative yields of fermentation products is3.89-fold higher than the produced by original strain. After six serial passages, dual-resistant mutant SG58relative yields of fermentation products remained stably, demonstrate its hereditary character is stably. Furthermore, the rpsL gene which encoding the ribosomal protein S12were analyzed. The gene sequencing result revealed that the rpsL gene sequence has not changed, in other words the corresponding amino acids sequence did not change, also. The expected results of this study to find the rpsL mutation did not occur.Phosphopantetheinyl transferases (PPTases) catalyze the essential post-translational activation of carrier proteins from fatty acid synthetases (FASs) in primary metabolism and polyketide synthetases (PKSs) and non-ribosomal polypeptide synthetases (NRPSs) in secondary metabolism. Phosphopantetheinylation occurs by transfer of the4’-phosphopantetheine (P-pant) prosthetic group from coenzyme A to a conserved serine residue in the carrier proteins, converting the proteins from their inactive "apo" forms to their active "holo" forms. Gene of phosphopantetheinyl transferase from Saccharopolyspora spinosa was cloned, expression and functional characterized in this work.