海洋细菌发酵产氨基寡糖素菌种筛选及条件选择研究
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摘要
采用几丁质选择性培养基富集和平板培养从北海红树林土样中分离出可降解几丁质产生透明水解圈的细菌12株,並从中筛选出一株可降解转化几丁质、产氨基寡糖较好的菌株(HB003,下同),经对其生物学特性和几丁质转化产氨基寡糖发酵试验,结果表明:该菌株的菌落圆形、湿润、浅黄;大小为0.5×1～2μm,革兰氏染色阴性,无芽孢,单根极生鞭毛,可运动;兼厌气,中温性,生长最适起始pH7.5～8.5,25～35℃,耐盐,16s rDNA序列分析显示,该菌株与弧菌(Vibrio)菌属中多株菌株相似性达99%。其在以几丁质为唯一碳源基质的液体培养基,起始pH 8.5,30℃的条件下静置发酵6 d,可产氨基寡糖153.5 mg/L;经对其产氨基寡糖素发酵条件的优化,试验结果表明:其在配方为:甲壳素10g、蛋白胨5 g、MgSO_4·7H_2O 0.6g、FeSO_4 0.01g、K_2HPO_4 0.9g、KH_2PO_4 0.6g、NaCl 20 g,蒸馏水1000 mL,pH 8.0的培养基,接种量1%(ν/ν)、在发酵温度30℃、通气量摇瓶转速为120 rpm的条件下发酵生产氨基寡糖素,产量可达到703.5 mg/L.
In this paper,adopting selective culture medium and brilliant clear circle were used to enrich and screen for marine bacteria strains producing chito-amino-oligosaccharides from the samples.12 strains were isolated.A strain with high chito-amino-oligosaccharides producing ability designated HB003 was chosen for studying its physiogical properties.It showed that the colony of HB003 is circular,moist and pale yellow in color,no spore,mobile by single polar flagellum,facultative anaerobic.It is Gram-negative.Its 16s rDNA sequence is highly similar with Vibrio Genus.The optimal initial pH for growth is 8～8.5 and the temperature is 25℃～30℃.It has NaCl resistance.153.5 mg/L chito-amino-oligosaccharides can be produced by fermentation of HB003 in initial pH 8.5 and 30℃chitin culture media for 6 days.The effects of compositions of culture medium and cultural conditions on chito-amino-oligosaccharids production were investigated.The optimal culture medium are: chitin 10 g,Peptone 5 g,MgSO_4·7H_2O 0.6 g,FeSO_4 0.01 g,K_2HPO_4 0.9 g, KH_2PO_4 0.6 g,NaCl 20 g,H_2O 1000 mL,pH 8.0;The optimal cultural conditions including fermentation temperature,rotation rate and amount of inoculation are:30℃,120 rpm,and 1%,respectively.The yield of chito-amino-oligosaccharides can be improved to 703.5 mg/L.
In this paper,adopting selective culture medium and brilliant clear circle were used to enrich and screen for marine bacteria strains producing chito-amino-oligosaccharides from the samples.12 strains were isolated.A strain with high chito-amino-oligosaccharides producing ability designated HB003 was chosen for studying its physiogical properties.It showed that the colony of HB003 is circular,moist and pale yellow in color,no spore,mobile by single polar flagellum,facultative anaerobic.It is Gram-negative.Its 16s rDNA sequence is highly similar with Vibrio Genus.The optimal initial pH for growth is 8～8.5 and the temperature is 25℃～30℃.It has NaCl resistance.153.5 mg/L chito-amino-oligosaccharides can be produced by fermentation of HB003 in initial pH 8.5 and 30℃chitin culture media for 6 days.The effects of compositions of culture medium and cultural conditions on chito-amino-oligosaccharids production were investigated.The optimal culture medium are: chitin 10 g,Peptone 5 g,MgSO_4·7H_2O 0.6 g,FeSO_4 0.01 g,K_2HPO_4 0.9 g, KH_2PO_4 0.6 g,NaCl 20 g,H_2O 1000 mL,pH 8.0;The optimal cultural conditions including fermentation temperature,rotation rate and amount of inoculation are:30℃,120 rpm,and 1%,respectively.The yield of chito-amino-oligosaccharides can be improved to 703.5 mg/L.
引文
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