背景:一种丝状真菌红青霉被广泛认可用于生产工业上重要的抗生素,工业规模的青霉素。
目的:为了更好地理解,野生型P.rubens的遗传蓝图是从印度分离的,以确定苯氧甲基青霉素的遗传/生物合成途径(青霉素V,PenV)和其他次级代谢产物。
方法:分离基因组DNA(gDNA),图书馆是按照Illumina平台准备的。根据IlluminaNovoSeq平台进行全基因组测序(WGS)。Further,SOAPdenovo用于组装由Bowtie-2和SAMtools软件包验证的短读数。使用Glimmer和GeneMark来挖掘基因组中的总基因。预测蛋白质的功能注释通过NCBI非冗余(NR)进行,UniProt,京都基因和基因组百科全书(KEGG),和基因本体论(GO)数据库。此外,通过SignalP4.1和TargetPv1.1以及通过CAZy数据库的碳水化合物活性酶(CAZymes)和蛋白酶家族进行分泌组分析。由Mauve2.4.0进行比较基因组分析。软件以发现P.rubensBIONCLP45和黄青霉威斯康星州54-1255之间的基因组相关性;还通过ParSNP工具用已知的青霉素产生菌株制备系统发育。
结果:红青霉菌BIONCLP45菌株从印度分离并产生过量的PenV。31.09Mb基因组以95.6%的覆盖率组装在参考基因组P.chrysogenumWis54-1255上,具有10687个蛋白质编码基因,3502个基因在NR中具有同源物,UniProt,KEGG,GO数据库此外,基因组中发现358个CAZymes和911个转运蛋白编码基因。基因组包含青霉素的完整途径,苯乙酸(PAA)分解代谢的匀浆途径,安德拉斯汀A,山梨醇,RoquefortineC,还有Meleagrin.BIONCLP45和Wis54-1255的比较基因组分析显示,2952个常见的KEGG直向同源蛋白质编码基因的覆盖率为99.89%。系统发育分析显示,BIONCLP45与Fleming的原始分离株P.rubensIMI15378成簇。
结论:该基因组可能是进一步研究高滴度青霉素生产的发酵过程和菌株工程方法的有用资源。
BACKGROUND: A filamentous fungus Penicillium rubens is widely recognized for producing industrially important antibiotic, penicillin at industrial scale.
OBJECTIVE: To better comprehend, the genetic blueprint of the wild-type P. rubens was isolated from India to identify the genetic/biosynthetic pathways for phenoxymethylpenicillin (penicillin V, PenV) and other secondary metabolites.
METHODS: Genomic DNA (gDNA) was isolated, and library was prepared as per Illumina platform. Whole genome sequencing (WGS) was performed according to Illumina NovoSeq platform. Further, SOAPdenovo was used to assemble the short reads validated by Bowtie-2 and SAMtools packages. Glimmer and GeneMark were used to dig out total genes in genome. Functional annotation of predicted proteins was performed by NCBI non-redundant (NR), UniProt, Kyoto Encyclopedia of Genes and Genomes (KEGG), and Gene Ontology (GO) databases. Moreover, secretome analysis was performed by SignalP 4.1 and TargetP v1.1 and carbohydrate-active enzymes (CAZymes) and protease families by CAZy database. Comparative genome analysis was performed by Mauve 2.4.0. software to find genomic correlation between P. rubens BIONCL P45 and Penicillium chrysogenum Wisconsin 54-1255; also phylogeny was prepared with known penicillin producing strains by ParSNP tool.
RESULTS: Penicillium rubens BIONCL P45 strain was isolated from India and is producing excess PenV. The 31.09 Mb genome was assembled with 95.6% coverage of the reference genome P. chrysogenum Wis 54-1255 with 10687 protein coding genes, 3502 genes had homologs in NR, UniProt, KEGG, and GO databases. Additionally, 358 CAZymes and 911 transporter coding genes were found in genome. Genome contains complete pathways for penicillin, homogentisate pathway of phenyl acetic acid (PAA) catabolism, Andrastin A, Sorbicillin, Roquefortine C, and Meleagrin. Comparative genome analysis of BIONCL P45 and Wis 54-1255 revealed 99.89% coverage with 2952 common KEGG orthologous protein-coding genes. Phylogenetic analysis revealed that BIONCL P45 was clustered with Fleming\'s original isolate P. rubens IMI 15378.
CONCLUSIONS: This genome can be a helpful resource for further research in developing fermentation processes and strain engineering approaches for high titer penicillin production.