| As an important model cyanobacteria,Synechocystis is an important research object in genetic engineering,metabolic engineering,and synthetic biology.It has broad application prospects in fields such as medicine,energy,chemical engineering,aquatic products,materials,and the environment.The genome of Synechocystis spp.has a large copy number and exhibits instability,which will have a certain impact on the research of genetic engineering,metabolic engineering,and synthetic biology.The key to solving this problem is to study the changes in its genome copy number and its regulatory mechanism.The genomic DNA of plant chloroplasts also has multiple copies.How chloroplast DNA(cp DNA)is maintained and inherited during development,and the impact of changes in cp DNA copy numbers on plant photosynthesis and plant cell metabolism are important research topics in plant physiology.Endosymbiosis theory believes that chloroplasts originate from blue algae.Therefore,studying the law of copy number changes and regulatory mechanisms of Synechocystis is also helpful for the study of the physiological functions of chloroplasts in green plants.Phosphorus plays a key role in the normal physiological and biochemical activities of cells,and is also an important component of DNA.Therefore,the content of phosphorus is directly related to the physiological processes and DNA content of Synechocystis cells.In this article,phosphorus stress was used as a culture condition to explore the changes pattern in the copy number of the genome of Synechocystis and the regulatory mechanism.The main research results were as follows:1.With phosphorus stress culture as the treatment group,and normal phosphorus culture as the control group,the alga was incubated under the same conditions for 30 days.Samples were taken every two days,and the content of the genome of Synechocystis was measured by fluorescence spectrophotometry,and the copy number of Synechocystis cells was measured by flow cytometry.The results showed that under phosphorus deficiency conditions,the growth rate of Synechocystis decreased;Under phosphorus containing culture conditions,the genomic content of Synechocystis spp.showed a characteristic of first increasing and then decreasing;Under phosphorus stress,the genome content of Synechocystis spp.was always lower than that of the control group,and had a downward trend;Flow cytometry analysis further verified the difference in the copy number of the genome of Synechocystis in the mid exponential period,and also found that there was a certain linear relationship between the number of copy numbers and the volume size of Synechocystis cells themselves.2.Transcriptome analysis was conducted on the metaphase of exponential growth of Synechocystis spp.under both phosphorus and phosphorus free culture conditions,including differential gene expression analysis,GO enrichment analysis,and KEGG enrichment analysis,and fluorescence quantitative PCR validation was performed on key genes.The results showed that genes in the dual component system,ABC transporters,purine metabolism,and pyrimidine metabolism were significantly upregulated under phosphorus deficiency conditions;Key genes in pathways related to DNA replication are significantly downregulated,and DNA helicase may be the key to controlling DNA copy number.3.The indicators related to photosynthesis(phycocyanin content,chlorophyll-a content),cell redox(thioredoxin peroxidase(TPX)activity),three major substances metabolism(total protein content),and nucleic acid metabolism(DNA endotomy activity,alkaline phosphatase activity)of Synechocystis spp.under both phosphorus and phosphorus free culture conditions were detected.The results showed that under phosphorus stress,TPX activity,phycocyanin content,chlorophyll-a content,alkaline phosphatase activity,and DNA endotomy activity were upregulated,while total protein content was downregulated.Conclusion: Phosphorus is an important factor affecting the genome copy number of Synechocystis.The number of copies is the result of coordination between DNA replication and degradation.Multicopy genomic DNA alleviates the need for phosphorus in cells under phosphorus deficiency conditions to some extent. |
