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2.2. Primary Screening of Marine Actinomycetes for Antimicrobial Activity The isolated marine actinomycetes were screened for their antimicrobial activity. The loopful of the growth from the slant was inoculated into 50 ml of seed medium; ASW-274(1) [glucose 15 g/L, corn steep liquor 5 g/L, sodium chloride 5 g/L, calcium carbonate 2 g/L, peptone 7.5 g/L, yeast extract 7.5 g/L, made in 75% ASW, pH 7.0 to 7.5] distributed in 250 ml Erlenmeyer flask and incubated for 96 h at 30˚C at 200 rpm on a rotary shakerdifferent production media used were as followed: ASW-SM12(1) [glucose 50 g/L, yeast extract 11 g/L, peptone 4 g/L beef extract 4 g/L, sodium chloride 2.5 g/L, calcium carbonate 5 g/L, made in 75% ASW, pH 7.4 -7.6; ASW-36P(1) [soluble starch 20 g/L, glucose 15 g/L, yeast extract 2 g/L, peptone 3 g/L, calcium carbonate 2 g/L, ammonium sulphate 0.5 g/L, corn steep liquor 2 g/L, sodium chloride 2 g/L, magnesium phosphate 5 g/L,cobalt chloride 0.001 g/L and TSS 1 ml/L (copper sulphate 7 g, ferrous sulphate 1 g, manganese chloride 8 g, zinc sulphate 2 g and demineralized water 1 L), made in 75% ASW pH 7.3 - 7.6]; ASW-1M [glycerol 30 g/L, glucose 3 g/L, yeast extract 2 g/L, sodium chloride 3 g/L, sodium nitrate 1 g/L, calcium carbonate 3 g/L, peptone3 g/L and TSS 1 ml/L, made in 75% ASW, pH 7.0 - 7.2]. The flasks were incubated for 96 h at 30˚C on rotary shaker at 200 rpm. The 1 ml of harvested whole broth was extracted with equal amount of methanol and centrifuged at 4000 rpm for 10 min. The supernatant was tested for antimicrobial activity by agar well diffusion method using different pathogenic strains [14]. LR grade chemicals were used in the study. |
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bxzc123: 金币+50, 翻译EPI+1, ★★★很有帮助 2014-11-01 12:46:28
bxzc123: 金币+50, 翻译EPI+1, ★★★很有帮助 2014-11-01 12:46:28
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2.2. 海洋放线菌的抗菌活性初筛 对分离的海洋放线菌进行筛选，选出其抗菌活性。用接种环将斜面培养基生长的菌接种到50ml种子培养基; 将ASW-274（1）[葡萄糖为15g /L，玉米浆5g /L，氯化钠5g/L，碳酸钙2g /L，蛋白胨7.5g /L，酵母提取物7.5g /L的，配制于75％ASW，pH为7.0〜7.5]，倒入250ml锥形烧瓶中，并在旋转摇床中，200rpm、30℃下温育96小时，使用下列不同的生产培养基：ASW-SM12（1）[葡萄糖为50g /L，酵母提取物11克/L，蛋白胨4克/升，牛肉提取物为4g /L，氯化钠为2.5g /L时，碳酸钙5g /L，配制于75％ASW，pH为7.4-7.6; ASW-36P（1）[可溶性淀粉20g /L，葡萄糖为15g /L，酵母提取物2克/升，蛋白胨3g /L，碳酸钙2g /L，硫酸铵0.5g /L，玉米浆2g /L，氯化钠2g /L，磷酸镁5g /L，氯化钴0.001克/L和TSS以1ml /L（硫酸铜7克，硫酸亚铁1克，氯化锰8克，硫酸锌2克和去离子水1升），配制于75％ASW，pH值7.3 - 7.6]; ASW-1M[甘油30g/L，葡萄糖3g /L，酵母提取物2克/升，氯化钠3g/L，硝酸钠1g/L，碳酸钙3g/L，蛋白胨3g/L和TSS以1ml /L，配制于75％ASW，pH值7.0 - 7.2]。烧瓶在旋转摇床以200rpm震荡、30℃下孵育96小时。用等量甲醇萃取1ml全培菌液，以4000rpm离心10分钟。琼脂扩散法测试上清液对不同致病菌株[14]的抗菌活性。本研究使用的化学品为LR级。 |
2楼2014-11-01 12:36:40
