Studies On The Methods Of Strain Improvement For Doramectin

Doramectin, C25-cyclohexyl-avermectin Bl, is one of the macrocyclic lactones antibiotics, which is produced by a Streptomyces avermitilis mutant HCB10042. To get high doramectin-producing strain, thee strain improvement models were studied including using rhodamine B, precursor and amino glucoside antibiotics as selective pressure on the plate, combined with UV, NTG, ⁶⁰Co or UV plus NTG as mutational factors. The mutant with about fifty times productivity higher than the parent strain was obtained.aveD, existing in the biosynthetic gene cluster of avermectins, is to encode C5-O-methyltransferase which catalyzes the formation of A component from B. The fomer has lower activity while the later has higher activity. In order to obtain a higher B component-producing mutant, we can disrupt aveD in the chomosome DNA of doramectin-producing strain with genetic manipulation. This work finished the PCR amplification of aveD, the sequence analysis of aveD, and constructed the plasmid of pLY34 carrying the sequence of disrupted aveD for homologous recombination with chomosome DNA of Streptomyces avermitilis. Second, the methods of protoplast transformation, electro-transformation and conjugation for foreign gene introduction were compared. At last, two transformants containing the plasmid of pLY34 were found by conjugation from E.coli.