牦牛溶菌酶的定向进化研究

发布时间：2018-06-01 20:00

本文选题：牦牛 + 溶菌酶　；参考：《黑龙江畜牧兽医》2017年09期

【摘要】：为了通过定向进化的方法提高牦牛溶菌酶的活性,试验选用3个牦牛基因YSL1a、YSL3c和YML,通过DNA洗牌技术进行序列重组以及酶活筛选,获得了一个重组基因HYL,检测其酶活,构建重组子基因HYL的真核表达载体,转化毕赤酵母X33,并对重组HYL溶菌酶的活性进行检测。结果表明:HYL与YSL1a、YSL3c和YML的序列相似性分别为85.36%、96.04%和83.11%,突变核苷酸分别为65,17,75个;重组序列与3个亲本基因相比均有序列缺失。筛选到一个酶活达667.72 U/mg的阳性克隆,其活性高于亲本基因中YSL3c真核表达产物。说明本试验中牦牛HYL的序列发生了序列重组,而且其真核表达产物具有活性。
[Abstract]:In order to improve the lysozyme activity of yak by directed evolution, three yak genes YSL1aYSL3c and YMLwere selected and sequenced and screened by DNA shuffling technique. A recombinant gene HYL was obtained and its enzyme activity was detected. The eukaryotic expression vector of recombinant gene HYL was constructed and transformed into Pichia pastoris X33. The lysozyme activity of recombinant HYL was detected. The results showed that the similarity of the sequence of 1: HYL with YSL1aHYSL3c and YML was 85.36% and 83.11%, respectively, and the mutation nucleotides were 65.17 and 75, respectively, and the sequence deletion of the recombinant gene was compared with that of the three parental genes. A positive clone with an enzyme activity of 667.72 U/mg was screened and its activity was higher than that of the eukaryotic expression product of YSL3c in the parent gene. The results showed that the sequence of yak HYL was recombined and its eukaryotic expression product was active.
【作者单位】：西南民族大学生命科学与技术学院;贵州省桐仁市农业委员会;
【基金】：四川省青年科技创新团队项目(2015TD0025) 科技部科技支撑项目(2014BAD13B03) 中央高校基本科研业务费专项资金项目(2014NZYQN59)
【分类号】：S823.85

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