| Myxobacteria are Gram-negative and unicellular gliding bacteria, which exhibit complicated multicellular social behavior. The microbes are capable of producing large numbers of bioactive compounds that show anticancer, antibacterial, fungicidal or immune-modulating activities.Myxococcus xanthus DK1622, Myxococcales order, Cystobacterineae suborder, Myxococcaceae class, Myxococcus genus, is the type strain for the study of myxobacteria social behavior and hereditary or evolution, which has single circle double-strand DNA but not endogenous plasmids, and its genome is about9.14Mb. The multicellular social behavior include dual motility, fruiting body formation and sporulation, predation and so on.Sorangium cellulosum, Myxococcales order, Sorangineae suborder, Polyangiaceae class, Sorangium genus, has the largest prokaryotic genome hitherto. As reported, almost half of these bioactive secondary metabolites of myxobacteria were identified from Sorangium, and nearly all of Sorangium strains could produce these bioactive compounds.Epothilones, which are anticancer mechanisms of paclitaxel(i.e. Microtubule stabilization) are produced by the Sorangium cellulosum. The polyketide compounds-epothilones has generated substantial interest over the last few years in the areas of chemistry, biology and medicine due to their interesting structure and more importantly, their activity against numerous cancer cell lines. Up to now, there are at least five epothilones or chemically modified derivatives of epothilone undergoing evaluation in clinical trials and one has already been authorized for clinical use and sale by the U.S. Food and Drug Administratio. However, the most serious problem is that the Sorangium cellulosum have poorly developed the genetic manipulation system and thus lacks flexible genetic tools. So the studies of epothilones and other secondary metabolites are lagging. To bypass the serious limitations, researchers integrated by homologous recombination the entire epothilones gene clusters to heterologous hosts, in which the background genetic methodology is already more highly developed, for the production of value metabolites. And Myxococcus xanthus DK1622is a suitable host that has the related substrate and enzyme for epothilones production.At present, structure modification and analogue synthesis of natural products are the main methods for screening new drugs. And new biosynthesis systems need to be expressed in suitable cell vectors, which have simple genomic structure-"minimum genome" to reduce noise and complication and to increase the controllability and maneuverability of systems. To make up the suitable vectors for new secondary metabolites synthesis, we begin to simplify myxococcus xanthusDK1622network via searching nonessential genes for deletion to form minimum genome.The research firstly predicted genomic islands (GIs) of DK1622using IslandPath tool, and there were19GIs whose length is about400kb,4.5%of DK1622genome sequence length. And we analyzed GIs’essential or not through gene annotation, protein blast, metabolism prediction and gene expression. The results show that>50%genes are hypothetical proteins, GIs genes do not to participate in basic metabolism network for growth, but part of them can express in growth or developmental stage. So we tried to knock out GI for its function analysis in growth. As a result, we got12mutant strains with GI deletion, which could grow normally. This work provide theoretical and experimental basis for minimum genome study.On the other hand, we studied the phenotype of12GI-deletion mutant strains. The results show that12GIs do not play a very important role in myxococcus xanthus development and predation, which would help us to understand myxobacteria evolution and adaptation better. |
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