Artificial cyanophages are considered to be an effective biological method to control harmful cyanobacterial bloom.However,no synthetic cyanophage genome has been constructed and where its obstacles are unclear.Here,w...Artificial cyanophages are considered to be an effective biological method to control harmful cyanobacterial bloom.However,no synthetic cyanophage genome has been constructed and where its obstacles are unclear.Here,we survey a stretch of 16 kb length sequence of cyanophage A-4L that is unclonable in Escherichia coli.We test 12 predicted promoters of cyanophage A-4L which were verified all active in E.coli.Next,we screen for eight ORFs that hindered the assembly of intermediate DNA fragments in E.coli and describe that seven ORFs in the 16 kb sequence could not be separately cloned in E.coli.All of unclonable ORFs in high-copy-number plasmid were successfully cloned using low-copy-number vector,suggesting that these ORFs were copy-number-dependent.We propose a clone strategy abandoned the promotor and the start codon that could be applied for unclonable ORFs.Last,we de novo synthesized and assembled the full-length genome of cyanophage A-4L.This work deepens the understanding of synthetic cyanophages studies.展开更多
Cyanobacteria,as essential photosynthetic microorganisms,play a pivotal role in aquatic ecosystems by contributing to primary production and nitrogen fixation.However,they can also lead to harmful cyanobacterial bloom...Cyanobacteria,as essential photosynthetic microorganisms,play a pivotal role in aquatic ecosystems by contributing to primary production and nitrogen fixation.However,they can also lead to harmful cyanobacterial blooms,threatening water quality and ecosystem stability.Cyanophages,viruses that specifically infect cyano-bacteria,are key regulators of cyanobacterial populations and drivers of microbial ecosystem dynamics.While marine cyanophage biology has been extensive studied,the mechanisms underlying freshwater cyanophage infections remain poorly understood.In this study,we investigate the infection dynamics of cyanophage A-4(L)in Nostoc sp.PCC 7120,a model species known for nitrogen fixation and cellular differentiation.Through in-tegrated transcriptomic and proteomic analysis,we revealed that A-4(L)infection is characterized by phased gene expression,with host transcription factors playing key regulatory roles.Notably,the infection induces downregulation of photosynthesis-related genes,likely reflecting a strategy to redirect host metabolic resources toward viral replication.Additionally,we examined how environmental factors,such as nitrogen sources and light conditions,influence these interactions.Our findings offer novel insights into phage-host dynamics in freshwater ecosystems,enhancing our understanding of phage-mediated ecological processes and paving the way for biotechnological and environmental applications.展开更多
基金supported by the grants from the National Key Research and Development Program of China (No.2018YFA0903000)the National Natural Science Foundation of China (31901019)and the China Postdoctoral Science foundation (2021M692389).
文摘Artificial cyanophages are considered to be an effective biological method to control harmful cyanobacterial bloom.However,no synthetic cyanophage genome has been constructed and where its obstacles are unclear.Here,we survey a stretch of 16 kb length sequence of cyanophage A-4L that is unclonable in Escherichia coli.We test 12 predicted promoters of cyanophage A-4L which were verified all active in E.coli.Next,we screen for eight ORFs that hindered the assembly of intermediate DNA fragments in E.coli and describe that seven ORFs in the 16 kb sequence could not be separately cloned in E.coli.All of unclonable ORFs in high-copy-number plasmid were successfully cloned using low-copy-number vector,suggesting that these ORFs were copy-number-dependent.We propose a clone strategy abandoned the promotor and the start codon that could be applied for unclonable ORFs.Last,we de novo synthesized and assembled the full-length genome of cyanophage A-4L.This work deepens the understanding of synthetic cyanophages studies.
基金supported by the National Key R&D Program of China(2018YFA0903100)Jiangxi Water Science and Technology Fund(202527ZDKT17).
文摘Cyanobacteria,as essential photosynthetic microorganisms,play a pivotal role in aquatic ecosystems by contributing to primary production and nitrogen fixation.However,they can also lead to harmful cyanobacterial blooms,threatening water quality and ecosystem stability.Cyanophages,viruses that specifically infect cyano-bacteria,are key regulators of cyanobacterial populations and drivers of microbial ecosystem dynamics.While marine cyanophage biology has been extensive studied,the mechanisms underlying freshwater cyanophage infections remain poorly understood.In this study,we investigate the infection dynamics of cyanophage A-4(L)in Nostoc sp.PCC 7120,a model species known for nitrogen fixation and cellular differentiation.Through in-tegrated transcriptomic and proteomic analysis,we revealed that A-4(L)infection is characterized by phased gene expression,with host transcription factors playing key regulatory roles.Notably,the infection induces downregulation of photosynthesis-related genes,likely reflecting a strategy to redirect host metabolic resources toward viral replication.Additionally,we examined how environmental factors,such as nitrogen sources and light conditions,influence these interactions.Our findings offer novel insights into phage-host dynamics in freshwater ecosystems,enhancing our understanding of phage-mediated ecological processes and paving the way for biotechnological and environmental applications.
