BACKGROUND

promoters have been used for single guide RNA (sgRNA) transcription in the clustered regularly interspaced short palindromic repeats/CRISPR-associated protein (CRISPR/Cas9) genome editing system. However, no available promoters have been identified in which is an important industrial platform for organic acid and protein production. Two CRISPR/Cas9 systems established in have recourse to the RNA polymerase II promoter or in vitro transcription for sgRNA synthesis, but these approaches generally increase cloning efforts and genetic manipulation. The validation of functional RNA polymerase II promoters is therefore an urgent need for .

RESULTS

Here, we developed a novel CRISPR/Cas9 system in for sgRNA expression, based on one endogenous promoter and two heterologous promoters. The three tested promoters enabled sgRNA transcription and the disruption of the polyketide synthase gene in . Furthermore, this system enabled highly efficient gene insertion at the targeted genome loci in using donor DNAs with homologous arms as short as 40-bp.

CONCLUSIONS

This study demonstrated that both heterologous and endogenous promoters were functional for sgRNA expression in . Based on this result, a novel and simple CRISPR/Cas9 toolbox was established in that will benefit future gene functional analysis and genome editing.