通过原生质体融合选育直接利用淀粉的肌苷产生菌

Selection of Inosine Producing Fusants By Protoplast Fusion

以产生ａ－淀粉酶的解淀粉芽孢杆菌（Ｂａｃｉｌｌｕｓａｍｙｌｏｌｉｑｕｅｆａｃｉｅｎｓ）ＷＤ－２１和不能利用淀粉的产生肌苷的枯草芽孢杆菌（ＢａｃｌｌｌｕｓＳｕｂｔｉｌｌｓ）ＨＷ－９（ａｄｅ，ＮＰ－，８－ＡＧ＇）为亲株，在分别以０．７５和０．５ｍｇ／ｍｌ的溶菌酶浓度作用下形成原生质体，在电镜观察下，其形成率为９９％。通过优化融合条件，即３５℃在ＰＥＧ－１０００溶液中融合，融合率达到３．１１×１０－６。得到的融合子在选择培养基上传代多次，淘汰掉不稳定的融合子，得到稳定的融合子Ｆ１６和Ｆ１２９具有产生ａ－淀粉酶的能力，在以淀粉为碳源的发酵培养基中摇瓶发酵试验，Ｆ１６菌株产生肌苷的能力接近６ｇ／Ｌ．

wo parent strains, Bacillus amyloliquefaciens WD-21 with erythromycin resistance producing a-amy-lase and Bacillus subtilis with 8-AGr resistance producing inosine, are fused by polyethylence glycoltreatment to form protoplast.The optimum condition for ptotoplast formation, regeneration and fusionhave been studied.The frequency of fusion is 3.11×10-6.The stable fusants are selected by 10 mg/L Ery and 100 mg/L 8-AGr medium plates. The fusants F16 obtained can preduce inosine in fermenta-tion broth in which starch is carbon source,the concentration of inosine is 5.89 g/L.