Ramie Degumming Strains By Protoplast Mutagenesis Breeding And The Research On Shake Flask Degumming Process

Due to high value of the comprehensive utilization,the ramie fiber is an important natural fiber resource.Because of its unique advantages,the ramie fiber is widely sold at home and abroad as high-grade textile raw materials.Containing glial components,ramie fibers can not be directly used in textile production,and it need have degumming treatment to meet textile requirements.Natural degumming method is subjective to en-vironmental conditions,time consuming,low production efficiency;enzymatic degumming production costs,not suitable for industrial production;chemical degumming method is to result in environmental pollution and detrimental to fiber quality.Natural bio-degumming,in a word,is a trend of development of the ramie degumming.In this thesis,we study the impact that the strain incubation time and lysozyme concentration,reaction temperature,hydrolysis time,the osmotic stabilizer have for the strain protoplast preparation and regeneration by using bacillus cereus B6 bacteria as theoriginal strain,saved by laboratory screening.Single factor test determine the range of factors,and incubation time and lysozyme concentration,reaction temperature,the four factors of enzymatic hydrolysis time orthogonal test.The results showed that:the strain liquid culture 8h,lysozyme concentration of 0.3mg/mL,hydrolysis temperature 32 ℃,hydrolysis time of 90min are optimal conditions for protoplast formation and regeneration.Using ultraviolet light and nitrosoguanidine(NTG) on the basis of protoplast formation and regeneration conditions,we have mutagenic treatment on strain protoplast and mortality and protoplast regeneration rate of the standard determine the best combined mutagenesis conditions.Screening of mutant strains by three rounds of mutagenesis,with enzyme activity and the rate of residual gum as a standard,finally a mutant(B6-7-26-31),the production of xylanase activity is 210.39U/mL,which is improved by about 110% than starting strain 100.33U/mL;the pectinase live production93.62U/mL is improved by about 65% than starting strain 56.85U/mL;residual gum content decrease from 9.1% to7.1%.Enzyme production capacity and residual gum content have little change for the six transfer,which demonstrate its genetic stability.Finally, through the research different carbon source, the nitrogen source, metallic ion and the ion concentration as well as each kind of rocker condition (fermentation time, temperature, rotation, vaccination quantity and initial pH) conditions for enzyme to sudden change of the influence B6-7-26-31, the determination best culture medium condition is: Sucrose 5‰, NH4NO3 1‰,protein peptone 1‰ and MgS04 · 7H2O 0.5‰,NaC10.1‰.The best flask fermentation degum condition:degum time 32h, degum temperature 35℃, rotation 180r/min, initial pH7.5, inoculum size7%.