高产L-乳酸米根霉的原生质体制备与再生条件研究  

Preparation and Regeneration of High-yield L-Lactic Acid-producing Protoplast from Rhizopus oryzae

作　　者：邱静[1] 罗水忠[1] 姜绍通[1] 郑志[1] 杨培周[1] 

机构地区：[1]合肥工业大学生物与食品工程学院,安徽合肥230009

出　　处：《食品科学》2011年第9期174-178,共5页Food Science

基　　金：国家"863"计划重点项目(2007AA10Z361);安徽省农产品精深加工重点实验室项目(2009AKSY0102);安徽省自然科学基金项目(090411015)

摘　　要：为建立原生质体融合法选育高产L-乳酸米根霉(Rhizopus oryzae)菌株,对米根霉菌株原生质体制备及再生条件进行研究。考察酶种类、酶质量浓度、时间、温度、pH值、渗透压、预处理等因素对原生质体制得量及再生率的影响,获得原生质体制备及其再生的优良条件:不添加甘氨酸,渗透压稳定剂为0.6mol/L的NaCl缓冲液,按1.5mg/mL组合酶质量浓度,蜗牛酶、纤维素酶和溶壁酶质量浓度配比为1:1:1进行混合,35℃、pH6、酶解2h,可获得原生质体制得量最大为2.74×106个/mL,再生率最高为6.8%。In order to construct a protoplast fusion method for the breeding of Rhizopus oryzae strains highly capable of producing L-lactic acid,Rhizopus oryzae mutant mut-a,which had been obtained in our laboratory,was used as parental strain for studying the preparation and regeneration of protoplast.The effects of enzyme type and loading,temperature,pH,osmotic pressure and pretreatment on protoplast yield and regeneration rate were probed,and their optimum levels were determined as follows： in the absence of glycin and the presence of 0.6 mol/L NaCl as osmotic pressure stabilizer,using a mixture（1.5 mg/mL total concentration） of snailase,cellulose and lywallzyme（1：1：1,m/m） for hydrolysis at 35 ℃ and pH 6 for 2 h.Under these conditions,the maximum protoplast yield and regeneration rate were 2.74 × 106 cells/mL and 6.8%,respectively.

关 键 词：L-乳酸 米根霉 原生质体 制备 再生 

分 类 号：Q935[生物学—微生物学]