Integrating Expression Of Vhb Gene In Streptomyces Bambergiensis And The Effect On The Biosynthesis Of Moenomycin

Streptomyces bambergiensis is a kind of strict areobacterium that can produce Moenomycin. The dissolved oxygen is one of the main limited factors that affected the accumulation of Moenomycin because of its mucous cultural broth.The study was carried out to promote synthesis of Moenomycin and improve cell growth by integrating expression of Vitreoscilla hemoglobin(vhb) gene in Streptomyces bambergiensis.Intergeneric transfer is an effective gene transfer method. It can avoid DNA's degradation by extracellular nuclease. What's more, the protocol is very easy .It has been applied in many streptomyces . Intergeneric transfer of Streptomyces bambergiensis is studied for the first time, and considered a very effective gene transfer tool.The promoter of resistance gene of erythromycin (ermEp) is a strong promoter which can constitutively express in streptomyces . Structure gene of vhb is placed under control of ermEp via SOE-PCR in order to express effectively .TGG codon of-35 section of ermEp,is mutated by deletion via OOE-PCR ,producting the stronger promoter ermE*p.pSET152 is a kind of integrating expression vectors. pBIB2004 and pBIB2005 are constructed by vhb gene is ligated to pSET152,and transferred to Streptomyces bambergiensis by E.coli-Streptomyces intergeneric transfer.lt is verified that vhb is successfully integrated to the chromosome by PCR,and the engineering strains express the Vitreoscilla hemoglobin with the bioactivity by CO binding difference spectrum and Western Blot.The studies have preliminary revealed that expression of Vitreoscilla hemoglobin may promote synthesis of Moenomycin and growth of the strain in the shaking flasks.The promotion played more important roles in more hypoxic conditions.We failed to get a significantly positive results in promotion of growth and production of antibiotics after exploring 20 litre fermentator.Perhaps It is concerned with the optimization of technological process. A further studies are needed.