联合囊肿
蓝藻
藻胆体
生物
发起人
光系统II
生物化学
光系统I
rpoN公司
光合作用
心理压抑
细胞生物学
化学
基因表达
基因
遗传学
突变体
细菌
作者
Deng Liu,Virginia M. Johnson,Himadri B. Pakrasi
标识
DOI:10.1021/acssynbio.0c00106
摘要
The cyanobacterium Synechocystis sp. PCC 6803 is used as a model organism to study photosynthesis, as it can utilize glucose as the sole carbon source to support its growth under heterotrophic conditions. CRISPR interference (CRISPRi) has been widely applied to repress the transcription of genes in a targeted manner in cyanobacteria. However, a robust and reversible induced CRISPRi system has not been explored in Synechocystis 6803 to knock down and recover the expression of a targeted gene. In this study, we built a tightly controlled chimeric promoter, PrhaBAD-RSW, in which a theophylline responsive riboswitch was integrated into a rhamnose-inducible promoter system. We applied this promoter to drive the expression of ddCpf1 (DNase-dead Cpf1 nuclease) in a CRISPRi system and chose the PSII reaction center gene psbD (D2 protein) to target for repression. psbD was specifically knocked down by over 95% of its native expression, leading to severely inhibited photosystem II activity and growth of Synechocystis 6803 under photoautotrophic conditions. Significantly, removal of the inducers rhamnose and theophylline reversed repression by CRISPRi. Expression of PsbD recovered following release of repression, coupled with increased photosystem II content and activity. This reversibly induced CRISPRi system in Synechocystis 6803 represents a new strategy for study of the biogenesis of photosynthetic complexes in cyanobacteria.
科研通智能强力驱动
Strongly Powered by AbleSci AI