**2.4. Orthologues of CRISPR/Cas9**

To date, the most common Cas9 protein used in plants is from the bacteria *Streptococcus pyrogens* (SpCAs9), which recognizes the NGG-type PAM [46]. Even though this PAM sequence is widely distributed across plant genomes, it does not cover the entire genome of the plant [46]. Many naturally occurring Cas9 orthologues impose distinct crRNA-tracrRNA duplex and PAM requirements [30]. Therefore, exploring orthologous Cas9 proteins with cognate gRNA and PAM sequences would greatly expand possible target sequences in a given genome and add new Cas9 orthologues with unique properties into the CRISPR/Cas arsenal [30]. As the alternative orthologous Cas9 requires different PAM sequences, the total number of possible target sites within a plant genome can be increased. Hence, the CRISPR/ Cas9 system is modified by cointroducing multiple Cas9 orthologue-based platforms with different effectors such as nuclease, transcription activator or repressor into the same cell, where they are guided by a specific group of gRNAs to carry out multiplex and complex manipulation of gene activities [30]
