Breeding Of Bacterial Cellulose High-yield Strains And Preliminary Study Of Mutation Effects

Bacterial cellulose is a kind of microbial synthesis of ultramicro pure cellulose.As a new style of biological material,compared with other nature of plant cellulose,it has better advantages in purity,crystallinity,water holding capacity,biocompatibility and biodegradable in nature.Now,it has been widely used in many areas,such as food,papermaking,medicine and biochemical engineering.It has a very wide range of commercial applications prospects and is a hotspot of today's biomaterials research.Until now,the application of bacterial cellulose is subject to certain restrictions because of its low production rate and high price,which as one of the bottlenecks of the cellulose industrial production and promotion of the application.Based on these problems,in order to improve the yield of bacterial cellulose and reduce production cost,this paper uses the DSE and 60Co-? to carry on compound mutagenesis on original strains,separates out the NaBr-NaBrO3 substratum,and analyze the gene differences of mutant strains which will be got later by using the technology of AFLP molecular markers,and reveals the direct and indirect contact between gene and bacterial cellulose production from the molecular level.And then,from the metabolic point of view,this paper analyzes the mutation's influences on bacterial cellulose biosythesis.Follows are the research results:We dealt with Gluconacetobacter hansenii HDMI-3 by DSE and 60Co-y compound mutagenesis,and finally we get three mutant strains.The mutant strains Br-3 obtained from primary mutagenic is 50%more than the bacterial cellulose production of original strains.The mutant strains Br-12 is low-yielding mutant strains,and its production reduced by 66.67%;bacterial cellulose production of high-yielding mutant strains which are got from compound mutagenesis of 60Co-? mutant strains Br-3 increases by 3.67 times.We use the AFLP technology to analyze the genetic differences between original strains and mutant strains.There appears three different fragments,the lost gene fragments of encoding TBDT proteins HD-bl will hinder the strains'utrient transport;the gene fragments HD-b2 of encoding extracellular polysaccharides export protein,it affects the extracellular release of bacterial cellulose bundle;the third different fragment HD-b3 is the gene sequences of unknown protein.There appears two different gene fragments,they are the acsD gene sequence HD-c2 and the lost of unknown function gene sequence HD-c2.acsD is one of the four operons of encoding cellulases,it affects the correct Exocellular bacterial cellulose's folding and assembly.Those different gene explains the reason of high yielding and low yielding of mutant strains to a certain extent.Based on the biosynthetic pathway of bacterial cellulose,in the analysis and direction of Extracellular metabolite,the production of mutant strains Br-12 yielded at the end of the intermediate metabolite gluconic acid fermentation is 1.48 times of that of M,3.28 times of Co-5.In the fermentation of citrate,the mutant strains Co-5's utilization rate is the highest,which is the 1.42%of origins,4.31%of mutant strains Br-12.Principal component analysis showed that H+ is the main factor for bacterial cellulose synthesis.Comprehensive analysis,the mutant strains in bacterial cellulose synthesis not only by the influence of genes on the cellulose synthesis pathway,but also from analysis of metabolites,to further more define the lower concentration of extracellular H+environment conducive to biosynthesis bacterial cellulose.