Ademakinwa Adedeji Nelson, Ayinla Zainab Adenike, Agboola Femi Kayode
Department of Biochemistry and Molecular Biology, Obafemi Awolowo University, Ile-Ife, Nigeria.
J Genet Eng Biotechnol. 2017 Dec;15(2):345-358. doi: 10.1016/j.jgeb.2017.06.012. Epub 2017 Jul 4.
Strain improvement of a low fructosyltransferase-producing NAC8 (Accession No. KX023301) was carried out using chemical mutagens such as ethidium bromide and ethyl methane sulfonate. The wild-type and mutant strain were distinguished using Random amplified polymorphic DNA PCR and DNA fingerprinting analysis. Plackett-Burman and Box Behnken design were statistical tools used to determine important media parameters and optimization, respectively. Phenotypically and genetically, the new improved strain was different from the wild-type. The most important media parameters from PDB influencing fructosyltransferase production were ammonium chloride, sucrose and yeast extract at < 0.05. Some significant parameters obtained with the BBD exhibited quadratic effects on FTase. The values (35.37 and 32.11), correlation coefficient (0.98 and 0.97) and the percent coefficient of variation (2.53% and 2.40%) were obtained for extracellular and intracellular FTase respectively. The validation of the model in the improved strain resulted in an overall 6.0 and 2.0-fold increase in extracellular and intracellular FTase respectively compared to the wild-type. A relatively low FTase-producing strain of NAC8 was enhanced for optimum production using a two-pronged approach involving mutagenesis and statistical optimization. The improved mutant strain also had remarkable biotechnological properties that make it a suitable alternative than the wild-type.
使用溴化乙锭和甲磺酸乙酯等化学诱变剂对低果糖基转移酶产生菌NAC8(登录号KX023301)进行菌株改良。通过随机扩增多态性DNA PCR和DNA指纹分析区分野生型和突变株。Plackett-Burman设计和Box Behnken设计分别是用于确定重要培养基参数和优化的统计工具。从表型和基因方面来看,新的改良菌株与野生型不同。对果糖基转移酶产生有影响的来自马铃薯葡萄糖肉汤(PDB)的最重要培养基参数是氯化铵、蔗糖和酵母提取物,P < 0.05。通过Box Behnken设计(BBD)获得的一些显著参数对果糖基转移酶表现出二次效应。胞外和胞内果糖基转移酶分别获得了值(35.37和32.11)、相关系数(0.98和0.97)以及变异系数百分比(2.53%和2.40%)。与野生型相比,改良菌株中模型的验证结果使胞外和胞内果糖基转移酶分别总体提高了6.0倍和2.0倍。采用诱变和统计优化相结合的双管齐下方法,提高了相对低产果糖基转移酶的NAC8菌株的产量以达到最佳生产水平。改良的突变株还具有显著的生物技术特性,使其成为比野生型更合适的替代菌株。
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