摘要
目的对耐热链霉菌S.thermotolerans ATCC 11416进行全基因组测序,并通过生物信息学软件对基因组信息进行分析,对其蕴藏的次级代谢产物生物合成基因簇(BGC)进行预测,并借助于比较基因组学等手段进行研究。方法培养S.thermotolerans ATCC 11416的菌丝体,提取基因组DNA,利用Illumina和PacBio平台进行全基因组测序。基因组序列通过anti-SMASH等预测次级代谢产物的BGC,推测其可能产生的化合物类型,利用定量qPCR初步判断目标BGC的表达情况,同时利用TBtools软件进行全基因组比对分析。结果S.thermotolerans ATCC 11416的基因组由1个线状染色体组成,基因组大小为8279432 bp,推测包含7356个基因,GC含量为71.65%,含有66个tRNA和18个rRNA序列,还鉴定出多种重复序列。从基因组中鉴定出26个次级代谢产物的BGC,其中Cluster 8是负责合成卡波霉素的BGC。利用qPCR检测并与Cluster 8中的关键基因表达情况比对发现Cluster 1、7、13、14、15、19和21中的功能基因表达活性较高,Cluster 5、6、9、11、12、16、18和24中的功能基因表达较低,而Cluster 2、3、4、10、17、20、22、23和25基因簇中的功能基因几乎不表达,为可能的沉默基因簇。Cluster 4、10、17、20、23和26与已知基因簇的同源性较低,很可能负责新化合物的合成。基因组比较分析发现,S.thermotolerans ATCC 11416与S.ambofaciens ATCC 23877具有较高同源性,两者都具有产生十六元环大环内酯类抗生素的潜力,在基因组比较中发现存在多处翻转、易位等基因组重排现象。结论链霉菌S.thermotolerans ATCC 11416具有中等基因组长度,包含了26个BGC,其中蕴含多种同源性低的沉默基因簇,为后续通过分子生物学手段激活这些沉默基因簇来获得新化合物奠定基础。
Objective Streptomyces thermotolerans is a producer of Carbomycin,which is commonly utilized in the industry for biotransforming Tylosin into acetylisovaleryltylosin.However,the whole genome sequence information of this strain has not been reported.Herein,the whole genome of S.thermotolerans ATCC 11416 was sequenced and analyzed by bioinformatics software to predict the putative biosynthetic gene cluster(BGC)of secondary metabolites,as well as comparative genomics research.Methods The mycelium of S.thermotolerans ATCC 11416 was cropped,the qualified genomic DNA was extracted,and the whole genome of S.thermotolerans ATCC 11416 was sequenced using sequencing platform Illumina combined with PacBio.The resultant genomic sequence was analyzed by anti-SMASH software to find the BGC of secondary metabolites and the deduced compounds produced by these BGC,and the transcription level of these BGC was identified by qPCR.The whole genome of ATCC 11416 comparisons with other streptomyces were analyzed by TBtools software.Results The genome of S.thermotolerans ATCC 11416 consisted of a linear chromosome with a genome size of 8279432 bp and was presumed to contain 7356 genes with a GC content of 71.65%.It contained 66 tRNA and 18 rRNA sequences,and multiple repeats sequences had been identified.The BGCs of 26 secondary metabolites were identified from the genome,of which Cluster 8 was responsible for carbomycin biosynthesis.Using qPCR detection and comparison with key genes in Cluster 8,it was found that the functional genes in Clusters 1,7,13,14,15,19 and 21 exhibited higher expression activity,while those in Clusters 5,6,9,11,12,16,18 and 24 showed lower expression activity.The functional genes in Clusters 2,3,4,10,17,20,22,23 and 25 were almost not expressed,indicating putative silencing gene clusters.Among the silent gene clusters,Clusters 4,10,17,20,23,and 26 were low homologous with the known gene clusters and might be responsible for synthesizing new compounds.Genomic comparative analysis showed that S.thermotolerans ATCC 11416 and S.ambofaciens ATCC 23877 exhibited higher homology,and both produced 16-membered macrolide antibiotics.Genomic rearrangement phenomena such as multiple flips and translocations were found in genomic comparison.Conclusion The thermotolerant actinomycete Streptomyces thermotolerans ATCC 11416 harbors a medium-sized genome containing 26 BGCs,including a number of silent gene clusters with low homology.This lays the foundation for the subsequent activation of these silent gene clusters through molecular biological approaches to obtain new compounds.
作者
任雅飞
江志波
赫卫清
郝天怡
REN Yafei;JIANG Zhibo;HE Weiqing;HAO Tianyi(Key Laboratory of Chemical Engineering and Technology of State Ethnic Affairs Commission,School of Chemistry and Chemical Engineering,Ningxia Low-grade Resource High Value Utilization and Environmental Chemical Integration Technology Innovation Team,North Minzu University,Yinchuan 750021,China;NHC Key Laboratory of Biotechnology of Antibiotics,CAMS Key Laboratory of Synthetic Biology for Drug Innovation,Institute of Medicinal Biotechnology,Chinese Academy of Medical Sciences&Peking Union Medical College,Beijing 100050,China)
出处
《中国医药生物技术》
2025年第3期241-252,共12页
Chinese Medicinal Biotechnology
基金
中央高校基本科研业务费(3332023048)。
