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采用酸性茚三酮法测定了 30株有益芽胞杆菌的赖氨酸产量 ,然后在不同的溶菌酶浓度下 ,对赖氨酸产量超过 0 .0 7g/ L 的 2 1株菌进行原生质体转化质粒 p UB110 ,测定原生质体形成率、原生质体再生率及转化频率 ,结果 6 10 3,6 10 4,6 12 0 ,6 12 9四株菌的转化频率较高。然后 ,采用经典遗传学方法选育AEC抗性突变株 ,使赖氨酸积累提高。其中 ,B.licheniformis 6 10 4诱变菌株 6 10 40 1能积累赖氨酸 2 .91g/L,比出发菌株提高了 17倍左右 ,转化率也提高了一个数量级。通过质粒的再转化试验及传代稳定性试验 ,进一步证实 B.licheniformis 6 10 4及其突变菌株 6 10 40 1是较好的受体菌 ,尤其是用于赖氨酸合成酶基因的表达。
Acidic ninhydrin method was used to measure the lysine yield of 30 strains of Bacillus licheniformis, and then the protoplast transformation plasmids of 21 strains with lysine production over 0.07 g / L under different concentration of lysozyme p UB110. The protoplast formation rate, protoplast regeneration rate and transformation frequency were determined. The results showed that the transformation frequencies of the four strains were higher than that of the other strains (6 10 3, 6 10 4, 6 12 0, 6 12 9). Then, the AEC-resistant mutant strain was bred by classical genetics to increase the accumulation of lysine. Among them, B.licheniformis 6 10 4 mutant strain 6 10 40 1 could accumulate lysine 2.991g / L, which was about 17 times higher than that of the original strain, and the conversion rate was also increased by one order of magnitude. Through the plasmid re-transformation test and passage stability test, it was further confirmed that B.licheniformis 6 10 4 and its mutant strain 6 10 40 1 are better recipient bacteria, especially for the expression of lysine synthase gene.