Table 2.

Tabular presentation of comparative attributes of plant genome editing techniques.

	CRISPR/Cas9	Zinc Finger Nucleases (ZFNs)	Transcription factor like effector nucleases (TALENs)	Reference
Mode of action	It works by inducing double-strand breaks in target DNA or single-strand DNA nicks (Cas9 nickase).	It can induce double-strand breaks in target DNA.	Induces DSBs in target DNA.	Li et al., 2013; Mao et al., 2013
Off target effects	These effects can be minimized by selecting unique crRNA sequence.	These have off-target effects.	Off target effects cannot be avoided.	Hsu et al., 2013
Generation of large scale libraries	YES, this is possible to generate large scale libraries.	Such generation is not possible because it requires customization of protein component for each gene.	Generation of large scale libraries is possible but technically difficult and challenging.	Cho et al., 2013; Hsu et al., 2013
Protein engineering steps	It does not requires protein engineering steps, very simple to test multiple gRNA.	It requires complex to test gRNA.	TALENs need protein engineering steps to test gRNA.	Cho et al., 2013
Cloning	Cloning is not necessary.	Cloning is necessary.	It requires cloning.	Cho et al., 2013
gRNA production	Any number of gRNA can be produced by in vitro transcription. It keeps budget away from extra load.	Bit difficult to produce this kind of RNA.	gRNA production is bit difficult to achieve through these effector nucleases.	Cho et al., 2013
Methylated DNA cleavage	It can cleave methylated DNA in human cells. This aspect is of special concern for plants as this has not been much explored	Unable to do so.	There are many question marks upon capacity of TALENs to perform methylated DNA cleavage.	Hsu et al., 2013; Ding et al., 2013
Multiplexing	This is main advantage of CRISPR. Several genes can be edited at same time. Only Cas9 needed	Highly difficult to achieve this through ZFNs.	Very difficult to obtain multiplexed genes by means of TALENs. Because it needs separate dimeric proteins specific for each target	Li et al., 2013; Mao et al., 2013
Structural proteins	CRISP R consists of single monomeric protein and chimeric RNA.	ZFNs work as dimeric and only protein component required.	TALENs also work as dimeric and require protein component.	Li et al., 2013; Upadhyay et al., 2013; Zhou et al., 2014
Catalytic domain	It has two cleavage domains called RUVC and HNH.	ZFNs have catalytic domain of restriction endonuclease FOKI which generates a DSB.	TALENs also have FOKI catalytic domain of restriction endonuclease for DSB generation.	Jinek et al., 2012
Mutation rate	Comparatively low mutation rate has been observed.	High mutation rate observed in plants.	Mutation rate is high as compared to CRISPR.	Li et al., 2013
Components	crRNA, Cas9 proteins	Zn-finger domains Non-specific FokI nuclease domain	Zn-finger domains Non-specific FokI nuclease domain	Kumar and Jain, 2014
Length of target sequence (bp)	20–22	18–24	24–59	Chen et al., 2016
Target recognition efficiency	High	High	High	Kumar and Jain, 2014
Level of experiment setup	Easy and very fast procedure of designing for new target site	Complicated procedure of redesigning for each new target site and need for expertise in protein engineering	Relatively easy procedure of designing for each new target site	Kumar and Jain, 2014