Type II CRISPR/Cas system has been divided in II-A, II-B, and II-C subtypes. The II-C subtype has two variants (II-C1, II-C2). The Type II system uses a single large multidomain Cas9 protein as the effector complex. Moreover, all Type II CRISPR/Cas loci contain cas1 and cas2 genes (essential for the CRISPR adaptation) and tracrRNA (noncoding RNA, required to mature the long pre-crRNA and for interactions with Cas9 protein). The crRNA-tracrRNA-Cas9 protein complex recognizes and cleaves the target DNA sequences. Cas9 nucleases recognize the PAM sequence (5ʹ-NGG-3ʹ) and induce blunt-end DSBs at target sites. Cas9 has two distinct nuclease domains, HNH and RuvC, which cleave the target and nontarget strand, respectively (Figure and Table ). Among the different types of CRISPR/Cas systems, the Type II system is the most widely used in genetic engineering due to its simplicity, versatility, and efficiency. The engineered CRISPR/Cas9 system encompasses two main components: a sgRNA and Cas9 protein. The sgRNA is a fusion of tracrRNA and crRNA, usually, 20 nucleotides long, complementary to the target sequence. The target DNA sequence should be adjacent to the PAM site in the genome. Inactivation of either nuclease domain of Cas9 creates a Cas9 nickase (nCas9), which cleaves only one DNA strand. Inactivation of both nuclease domains generates dead Cas9 (dCas9), which still binds to target DNA. nCas9 is useful in base editors and prime editors, which perform precise genome editing without requiring DSBs, and dCas9 serves as a scaffold for recruiting effectors proximal to specific genomic sites. dCas9 is widely used for regulating transcription, altering epigenetic controls, imaging living cells, and other purposes. Type V CRISPR/Cas system can be divided into V-A to V-I, V-K, and V-U subtypes with diverse functions and its effectors are Cas12a (Cpf1), Cas12b (C2c1), Cas12c (C2c3), etc. Type V CRISPR/Cas system requires Cas and crRNA to edit the target site. Once the crRNA recognizes the PAM site (5'-TTTN-3') and fully pairs with the target DNA base, Cas12a uses the RuvC domain to cut the target sequence in a cis manner to generate the 5' sticky end, and at the same time, uses the trans cutting activity to cut any adjacent ssDNA. Cas12a shows RNase III activity, does not require a tracrRNA, and generates mature crRNA. Like Cas12a, Cas14a belongs to the Class 2 system Type V family. Cas14a proteins have RNA-guided ssDNA-targeted endonuclease activity. Cas14a proteins are not required to recognize PAM sites in the DNA sequence. Type VI CRISPR/Cas system can be divided into VI-A, VI-B, VI-C, and VI-D subtypes. The signature protein is Cas13 (C2c2), which has higher eukaryotes and prokaryotes nucleotidebinding (HEPN) domains. The uniqueness of this system is that Cas13 can recognize ssRNA molecular targets. Under the targeting effect of crRNA without the requirement of tracrRNA, the Cas13-crRNA complex recognizes the sequence of the protospacer flanking site (PFS) on the target nucleic acid, and at the same time cuts the target RNA, trans cuts the single-strand RNA (ssRNA).