Systems metabolic engineering for citric acid production by Aspergillus niger in the post-genomic era

Table 2 CRISPR/Cas9 genome editing systems used in A. niger

sgRNA expression		Gene editing	Donor DNA (SM/Homology arm size, bp)	Efficiency	Advantages	Limitation	References
Promoter	Terminator	Gene editing	Donor DNA (SM/Homology arm size, bp)	Efficiency	Advantages	Limitation	References
PgpdA	TtrpC	NHEJ-mediated gene disruption	–	Some	Less PAM limitation	Requiring to add HH and HDV, more cloning effort	[84]
In vitro synthesis		HR-mediated gene deletion	pyrG/1500	28–100%	Instantaneous genome editing	Hard to be used in gene regulation, dependent on sgRNA uptake and stability	[85]
PmbfA	TtrpC	NHEJ-mediated gene disruption	–	Some	Less PAM limitation	Requiring to add HH and HDV, more cloning effort	[87]
PmbfA	TtrpC	HR-mediated gene integration	pyrG/690 and 834	100%	Less PAM limitation	Requiring to add HH and HDV, more cloning effort	[87]
PhU6	Ploy (T)₆	NHEJ-mediated gene disruption	–	15%	Less cloning effort	PAM motif (GN₁₉GG)	[88]
PyU6			–	20%
PanU6			–	23%
PanU6		HR-mediated gene integration	hph/40	36%
5S rRNA	Ploy (T)₆	NHEJ-mediated gene disruption	–	95.28–100%	High efficiency, less cloning, less PAM limitation	–	[89]
		HR-mediated gene integration and deletion	hph/40	100%
		Multiplex gene editing	hph/40	45.83%

ISSN: 1475-2859