论文部分内容阅读
【目的】高质量的海洋微生物菌种库及其天然产物库是新药开发的重要来源。在本研究中我们通过筛选新的烯二炔类抗生素来对已经构建的海洋微生物天然产物库进行质量评价。【方法】首先我们根据烯二炔类抗生素能引起DNA断裂的活性构建了活性筛选模型,并对我们的天然产物库进行筛选;其次根据合成烯二炔核心结构的独特且保守的重复I型聚酮合成酶设计了引物,通过PCR扩增的方法对海洋微生物库进行序列筛选。【结果】通过活性筛选从我们的海洋微生物天然产物库中获得一个阳性的发酵产物。对该阳性菌(LS481)的系统发育学分析表明该菌属于能产生烯二炔类化合物—Dynemicin的Micromonospora chersina,对其发酵产物TLC分析证明该菌确实产生Dynemicin类化合物。通过基因筛选得到了2个具备合成烯二炔核心结构聚酮合成酶的菌株,16S rRNA基因分析显示其中一个很可能为灰色链霉菌(MS098),另外一株菌则同Streptomyces vinaceus NBRC 13425T和Streptomyces cirratus NRRLB-3250T最相近。【结论】我们的活性筛选模型能够有效获得烯二炔类物质,结合基因筛选能够进一步获得可能产生烯二炔物质的菌株。初筛结果也再一次验证了我们海洋微生物天然产物库的质量较好。
【Objective】 High-quality marine microbial species pool and its natural product pool are important sources of new drug development. In this study, we evaluated the quality of the marine natural microbial pools that have been constructed by screening new enediyne antibiotics. 【Method】 Firstly, we constructed an active screening model based on the activity of enediyne antibiotics that causes DNA cleavage and screened our natural product library. Secondly, based on the unique and conservative repeat type I polyketide Keto-synthetase primers were designed and the library of marine microorganisms was screened by PCR amplification. [Results] A positive fermentation product was obtained from our marine microbial natural product pool by active screening. Phylogenetic analysis of this positive bacterium (LS481) showed that this strain belonged to Micromonospora chersina, which produces the enediyne-Dynemicin. TLC analysis of its fermentation product proved that the strain did produce Dynemicin. Two strains of polyketide synthases with core structure of enketidene were obtained through gene screening. One of the 16S rRNA gene analyzes is likely to be Streptomyces griseus (MS098) and the other strain is identical to Streptomyces vinaceus NBRC 13425T and Streptomyces cirratus NRRLB-3250T most similar. 【Conclusion】 Our active screening model can effectively obtain enediynes, and the combination of gene screening can further obtain strains that may produce enediyne. The results of the preliminary screening once again verify that the quality of our marine microbial natural product library is better.