Medires Publishers - Article

Abstract

Genome editing offers a range of solutions for more efficient development of crops that are productive, adapted to stresses, climate-resilient, and less dependent on agro-inputs. Clustered regularly interspaced short palindromic repeats (CRISPR)–CRISPR-associated protein (Cas9) technology is the current dominant tool used for genome editing. Originally, a Cas9 nuclease was employed to induce a double-strand break in its target site, causing the deletion of a few base pairs, inversion, and gene integration to deliver desired changes in organisms. Aside from the primary nuclease activity (knock-in/out), a Base editor system, Gene priming and Cargo chauffeuring activities have been reported to deliver functionalities to specific regions in the DNA such as regulating transcription and fluorescence DNA for visualizing and understanding biological systems. Limitations of the scope of Cas9 activity were also eliminated by the recent development of more Cas9 orthologues (Cpf1-RR and Cpf1-RVR). Cas9 together with the advent of novel base editing tools that enable precise genome modifications and DNA-free genome editing via ribonucleoproteins, demonstrates significant promise in the development of future crop improvement strategies. However, large-scale adoption of CRISPR/CAS will require optimizing strategies while accounting for costs, ease of implementation, and potential impacts on production gains. This review focuses on CRISPR application in plants, advances in CRISPR technology, regulations that may disadvantage scientists, resources for the smooth application of CRISPR and the preparedness of Africa to benefit from CRISPR technology.

Introduction

Mutagenesis conferring genome changes in organisms may either turn off (Gene knockouts) or on (Gene knock-in) the function of genes in target regions. This results from a disruption in the synthesis of essential amino acids relevant in diverse metabolic pathways. Genetic mutants are essential for elucidating the genetic and molecular controls of many important biological mechanisms (Fang et al., 2018; Li et al., 2018; Ding et al., 2016; Matsumoto, 2005). The use of mutant-induced genetic variations in plant breeding commenced in the 1960s when radiation and chemical mutagenesis were developed to select favorable genetic combinations. Alterations in the sequence of the Deoxyribonucleic acid (DNA) (single or double-stranded) of organisms are critical for gene disruption (Durland & Ahmadian-Moghadam, 2021; Chaud- hary et al., 2019). At present, gene transfer and elimination can be achieved through conventional crosses, mutagenesis or through biotechnological approaches like genome editing(Sedlar, 2020; Cardi, 2016). Mutant libraries of several model plants have been generated by physical, chemical, or insertion (T-DNA/transposon insertion) mutagenesis and, more recently,by genome editing techniques (Lu et al., 2017; Meng et al. 2017). Traditional mutagenesis approaches often generate random mutations in the host genome, which could often cause deleterious effects that may not be intended and have many drawbacks including large screening populations to identify desired variants (Mohanta et al., 2017; Sikora et al., 2012; El-Gewely et al., 2005). Since the late 2000s four families of engineered and programmable nucleases including Me- ga-Nucleases, Zinc Finger Nucleases (ZFNs), Transcription Activator-Like Effector nucleases (TALEN) and the Clustered Regularly Inter-Spaced Short Palindromic Repeats (CRIS- PR) Cas9 changed mutagenesis from random events to tar- geted modification of genomes (Mohanta et al., 2017; Gaj et al., 2016; Liang et al., 2014). These developments have limited mutations to specified regions of the genome, which, have helped decipher the function of genes more accurately. Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas9-associated protein technology is the cur- rent dominant genome editing tool used to manipulate and change the DNA of organisms (Ahmad et al., 2020;Wada et al., 2020;Tripathi et al., 2020;Li et al., 2017; Haeussler and Concordet, 2016). The CRISPR technology mimics the sur- veillance system performed by immune memory in bacteria that enables it to recognize and degrade foreign nucleic ac- ids (Koonin et al., 2017; Doudna and Charpentier, 2014;Fin- eran and Charpentier, 2012). The engineered CRISPR-Cas9 system relies on single guide RNA (sgRNA), a small non-coding RNA composed of a crR- NA homologous to a genome target region and a tracrRNA that binds to CAS9. Target sites are generally located in the exon regions of an open reading frame in order to induce effective frameshift mutations (Jinek et al., 2012; Biswas et al., 2019).The target region of CRISPR sgRNA is usually 3 bp upstream of the NGG Proto-spacer Adjacent Motif (PAM) (Meng et al., 2017). The Cas9 has a bi-lobed architecture, with a large globu- lar recognition lobe (REC) and a small nuclease lobe (NUC) with two nuclease domains, RuvC and HNH, used to edit the DNA at the right position (Li et al., 2018; Endo et al., 2018;Lei et al., 2014;Voytas, 2013) The CRISPR/Cas9 system can be delivered to targeted cells either indirectly by biological transfer (Agrobacterium tumefaciens) or directly by microprojectile bombardment of cul- tured tissues, electroporation, or chemical treatment (polyethylene glycol—PEG) of protoplast suspensions. Alternatively, it can be delivered as ribonucleoprotein complexes in bom- barded tissues or protoplasts (Sharma et al., 2020;Liang et al., 2019;Murovec et al., 2018; Liang et al, 2017; Ding et al., 2016; Doench et al., 2016; Zhang et al., 2016;Krenek et al., 2015;Woo et al., 2015; Mao et al., 2013). However, CRISPR applications in plants have largely depended on Agrobacterium-mediated T-DNA transformation, which, is limited to a narrow range of genotypes within a species. The traditional CRISPR genome editing process relies on the double-strand break (DSB) repair capacity of recipient cell types, the promoter under which the Cas9/gRNA is expressed and the regeneration efficiency of the crop (Ding et al., 2016). The DNA breaks caused by the nuclease are generally repaired by non-homologous end joining (NHEJ) eventually causing deletion and/or insertion of nucleotide(s) or, when a DNA repair template with homology regions with the target site is provided, homology-directed repair (HDR) (Lieber, 2010) (Figure 1). In the NHEJ approach, the cell repair mechanism attempts to rejoin the cut ends but loses or induces a few bases in the process: a situation that can result in gene alterations due to frameshift mutations. In the HDR situation, donor repair templates are supplied, and the DSB repair may results in the integration of the donor sequence to allow targeted modification of genes by small to large nucleotide replacement or insertions, but its efficiency remains low in flowering plants (Endo et al., 2018; Li et al., 2018; Shibata, 2017; Jiang et al., 2017;Zhang et al., 2017;Gaj et al., 2016; San Filippo et al., 2008).

Figure 1: Repair of double strand break in DNA (Adapted from Wu et al., 2020) Advances in CRISPR-Cas9 technology development and applications The original CRISPR-Cas9 protein uses a combination of an enzyme that cuts DNA (Cas9, a nuclease) and a guiding piece of genetic material (guide RNA) that specifies the location in the genome to be targeted. The application scope of CRISPR – Cas9 was expanded with the use of new CAS9 variants or CAS proteins with different specificities, notably the PAM, base editing, prime editing and simultaneously targeted mutations (multiplexing) (Wu et al., 2020; Wang et al., 2018; Ding et al., 2016). Recently, Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-Cas12a nuclease (Cpf1), dCas9 transcriptional regulators, base editors, PRIME editors, and RNA editing tools are widely used in basic research (Wu et al., 2020; Ai, et al., 2022).

Recognition of Guide RNA

The original CRISPR-Cas9 technology is based on the commonly used Streptococcus pyogenes Cas9 system which recognizes only Proto-spacer adjacent motif (PAM) = NGG. Alternative genome editing technologies including CRISPR Prevotella and Francisella 1 (Cpf1) or Cas12a have opened up CRISPR editing to additional areas of the genome (Shmakov et al.,2017., Gao et al., 2017). The Cas9 and Cpf1 differ in the structural organization as the Cpf1 protein is smaller, use of a shorter guide as the matu- ration of crRNA by Cpf1 does not require the assistance of trans-activating crRNA (tracrRNA). Whereas Cas9 cleaves the DNA close to the PAM site with blunt ends, Cpf1 cleaves distal to the PAM site and produces staggered ends, which offer the opportunity for further modification at the same would also facilitate HDR. The Cpf1 recognizes TTTV as well as TTTT PAM sites increasing the target range for ge- nome editing for the Cas9 tool (Jiang et al., 2017; Ding et al., 2016; Kleinstiver et al., 2015). The Cas9 and Cas12a both cut DNA, while the Cas13 family has been shown to target RNA (Ai et al. 2022; Tang et al. 2021; Yangs et al. 2019)

Nuclease Activity

Recently, new and more powerful techniques to edit genes have been developed based on the disabled activity of the Cas9 nuclease. When the nuclease activity of the Cas9 protein is disabled, it is referred to as dead (dCas9). The dead Cas9 can be fused to transcription activators, epigenetic modifiers and Fluorescence-labeled Epitope-tagged to modulate transcription reporter gene function (Veillet et al., 2020; Li et al., 2016; Kleinstiver et al., 2015; Fu et al., Multiplex genome editing). Another important development in the CRISPR-Cas9 technology is the ability to effectively express multiple sgRNAs for simultaneous editing of two or more 2014; Mercer et al., 2012). New technologies have been developed for gene activation, replacement, and insertion to satisfy various demands of genome editing for functional genomic studies, which eliminate the necessary injury due to the double-strand break (DSB) associated with traditional Cas9 technology.

Base Editing

Is the conversion of one base or base pair into another (e.g., A:T to G:C, C:G to T:A) or the introduction of point mutations at specific sites which permits the recoding of the DNA sequences (Li et al., 2021; Veillet et al., 2020; Li et al., 2018; Kim et al., 2017; Zong et al., 2017). The Adenine (Adenine - Guanine) and cytidine (cytosine-uracil-thiamine) base editors introduce mutations that largely do not disrupt the function of genes (Kim et al., 2017). The cytidine deaminase enzyme removes an amino group from cytosine, converting it to uracil, resulting in a U-G mismatch, which gets resolved via DNA repair pathways to form U-A base pairs. Subsequently, a T gets incorporated into the newly synthesized strand, forming T-A base pairs. This results in C-G to T-A conversion in a programmable manner. Unlike cytidine deaminases, adenine DNA deaminases do not occur in nature. In 2017, David Liu and the group developed ABEs by using Escherichia coli TadA (E. coli TadA) through extensive protein engineering and directed evolution. Escherichia coli TadA is a tRNA adenine deaminase that converts adenine to inosine in the single-stranded anticodon loop of tRNA Arg. It shares homology with the APOBEC enzyme. The procedure involves the conversion of Cas9 nuclease to nickase followed by the fusion of the nickase with the deaminase enzyme (either cytidine deaminase or engineered adenine deaminase for C-G to A-T and A- T to C-G conversions respectively) (Veillet et al., 2020). The base editing technique permits the conversion of alleles by base change, the creation of premature stop codon for knockout, and the creation of new alleles of genes that do not exist in nature and has the advantage of overcoming the long procedure of introgressing new alleles. For instance, the acetolactate synthase (ALS) gene in tomatoes has been edited using Agrobacterium-mediated transformation with a Cytidine base editor changing amino acids essential for the protein to be targeted by the chlorsulfuron herbicide (Veillet et al., 2019). The authors reported the production of chlorsulfuron-tolerant plants with up to 71 genes (Wang et al., 2018;Ma et al., 2015; Lowder, et al., 2015; Cong et al., 2013; Kurata, et al., 2018; Li et al., 2018; Wang et al., 2017). Multiplexing thus addresses the extremely time-consuming and laborious bottleneck of conventional cross-breeding methods for pyramiding multiple QTLs (Abdelraman et al., 2021). In China, a CRISPR/Cas9-mediated multiplex genome editing system was used to simultaneously mutate three major genes in rice which negatively regulated grain weight including Grain Width 2 (GW2), Grain Width 5 (GW5) and Thousand-Grain Weight 6 (TGW6)(Dai et al., 2016). All three genes generated mutants (gw5tgw6 and gw2gw5tgw6) which showed notably larger grain sizes than that of the wild-type (Dai et al., 2016). In hexaploid bread wheat, CRISPR-Cas9 has been used to modify multiple alleles to confer heritable resistance to powdery mildew (Wang et al. 2014b, Wang et al. 2018).

Transgene free Edited Products

An important feature of the agricultural application of CRISPR-Cas9 technology is the delivery of nontransgenic traits in crops. In seed-propagated crops, the CRISPR/Cas9 transgene can be easily eliminated through Mendelian segregation in progenies. This process alone distinguishes the technique from the class of GMOs and hence absolves CRISPR edits from rigorous testing regimes. The transformation of protoplasts followed by plant regeneration allows either the delivery of a ribonucleoprotein complex or the transient expression of delivered plasmid DNA without DNA integration into the host genome. Such methods are particularly useful for vegetatively propagated crops. Woo et al. (2015) have optimized CRISPR/Cas9 ribonucleoproteins (RNPs) to completely avoid transgene integration. Veillet et al. (2019) reported 12.9% and 10 Applications of CRISPR technologies in crop improvement. 3.1 CRISPR-Cas9 genome editing methods have been exploited in several spheres of research including breeding and development of agricultural crops, animals, and human health genetic applications (Ahmad et al., 2020; Tripathi et al., 2020; Wada et al., 2020; Bao et al., 2019; Ding et al., 2016). In agriculture, CRISPR-Cas9 has been applied to modify and understand the functions of genes controlling major agronomic, nutritional and economic traits in a vast array of crops to achieve improved yield performance, enhanced nutritional quality (biofortification), and biotic and abiotic stress tolerance (Johnsson et al., 2019; Romero and Gatica-Arias, 2019; Mishra and Zhao, 2018; Mohanta et al., 2017; Zhang et al., 2017). There are several efforts of gene function analysis in important crops including bananas, cassava, maize, millet rice, sorghum, soybeans, tobacco, and tomatoes. Table 1 shows some of the current applications of CRISPR/Cas9 systems in crops are for studying gene function, improving agricultural traits, creating male-sterility mutants, and inducing haploids. The role of genomics in plant research and sustainable crop production is imminent, and it will become even more crucial in the future. Two key applications of CRISPR that will revolutionize agriculture more than the green revolution are the fixing of hybrids and the enhancing of diversity to improve adaptation.

Promoting apomixis for hybrid vigor preservation through the CRISPR/Cas9 editing

References

1. Abdelrahman, Mohamed, Zheng Wei, Jai S. Rohila, and Kaijun Zhao. "Multiplex genome-editing technologies for revolution- izing plant biology and crop improvement." Frontiers in Plant Science 12 (2021): 721203.
   View at Publisher | View at Google Scholar
2. Ahmad, Shakeel, Xiangjin Wei, Zhonghua Sheng, Peisong Hu, and Shaoqing Tang. "CRISPR/Cas9 for development of disease resistance in plants: recent progress, limitations and future prospects." Briefings in Functional Genomics 19, no. 1 (2020): 26-39.
   View at Publisher | View at Google Scholar
3. Ai, Yuxi, Dongming Liang, and Jeremy E. Wilusz. "CRISPR/ Cas13 effectors have differing extents of off-target effects that limit their utility in eukaryotic cells." Nucleic Acids Research 50, no. 11 (2022): e65-e65.
   View at Publisher | View at Google Scholar
4. Anders, Carolin, Katja Bargsten, and Martin Jinek. "Structur- al plasticity of PAM recognition by engineered variants of the RNA-guided endonuclease Cas9." Molecular cell 61, no. 6 (2016): 895-902.
   View at Publisher | View at Google Scholar
5. Bao, Aili, David J. Burritt, Haifeng Chen, Xinan Zhou, Dong Cao, and Lam-Son Phan Tran. "The CRISPR/Cas9 system and its applications in crop genome editing." Critical reviews in biotechnology 39, no. 3 (2019): 321-336.
   View at Publisher | View at Google Scholar
6. Barcaccia, Gianni, and Emidio Albertini. "Apomixis in plant re- production: a novel perspective on an old dilemma." Plant re- production 26 (2013): 159-179.
   View at Publisher | View at Google Scholar
7. Biswas, Sukumar, Rong Li, Zheng Yuan, Dabing Zhang, Xiangxiang Zhao, and Jianxin Shi. "Development of methods for effective identification of CRISPR/Cas9-induced indels in rice." Plant Cell Reports 38 (2019): 503-510.
   View at Publisher | View at Google Scholar
8. Blary, Aurélien, and Eric Jenczewski. "Manipulation of cross- over frequency and distribution for plant breeding." Theoretical and Applied Genetics 132 (2019): 575-592.
   View at Publisher | View at Google Scholar
9. Nakato, Valentine. "Guy Blomme, Kim Jacobsen, Walter Oci- mati, Fen Beed, Jules Ntamwira, Charles Sivirihauma, Fred Ssekiwoko." Eur J Plant Pathol 139 (2014): 265-281.
   View at Publisher | View at Google Scholar
10. Callaway, Ewen. "CRISPR plants now subject to tough GM laws in European Union." Nature 560, no. 7716 (2018): 16-17.
    View at Publisher | View at Google Scholar
11. Cardi, Teodoro. "Cisgenesis and genome editing: Combining concepts and efforts for a smarter use of genetic resources in crop breeding." Plant Breeding 135, no. 2 (2016): 139-147.
    View at Publisher | View at Google Scholar
12. Chakraborty, M., P. Sairam Reddy, M. Laxmi Narasu, Gaurav Krishna, and Debashis Rana. "Agrobacterium-mediated genet- ic transformation of commercially elite rice restorer line using npt II gene as a plant selection marker." Physiology and molec- ular biology of plants 22 (2016): 51-60.
    View at Publisher | View at Google Scholar
13. Chaudhary, Juhi, Rupesh Deshmukh, and Humira Sonah. "Mutagenesis approaches and their role in crop improvement." Plants 8, no. 11 (2019): 467.
    View at Publisher | View at Google Scholar
14. Chelysheva, Liudmila, Stéphanie Diallo, Daniel Vezon, Ghis- laine Gendrot, Nathalie Vrielynck, Katia Belcram, Nathalie Rocques et al. "AtREC8 and AtSCC3 are essential to the mo- nopolar orientation of the kinetochores during meiosis." Jour- nal of cell science 118, no. 20 (2005): 4621-4632.
    View at Publisher | View at Google Scholar
15. Cai, Yupeng, Li Chen, Xiujie Liu, Chen Guo, Shi Sun, Cunxiang Wu, Bingjun Jiang, Tianfu Han, and Wensheng Hou. "CRISPR/ Cas9-mediated targeted mutagenesis of GmFT2a delays flow- ering time in soya bean." Plant biotechnology journal 16, no. 1 (2018): 176-185.
    View at Publisher | View at Google Scholar
16. Claeys, Hannes, Son Lang Vi, Xiaosa Xu, Namiko Satoh-Na- gasawa, Andrea L. Eveland, Alexander Goldshmidt, Regi- na Feil et al. "Control of meristem determinacy by trehalose 6-phosphate phosphatases is uncoupled from enzymatic activ- ity." Nature plants 5, no. 4 (2019): 352-357.
    View at Publisher | View at Google Scholar
17. Cong, Le, F. Ann Ran, David Cox, Shuailiang Lin, Robert Bar- retto, Naomi Habib, Patrick D. Hsu et al. "Multiplex genome engineering using CRISPR/Cas systems." Science 339, no. 6121 (2013): 819-823.
    View at Publisher | View at Google Scholar
18. Dai, Wei-Jing, Li-Yao Zhu, Zhong-Yi Yan, Yong Xu, Qi-Long Wang, and Xiao-Jie Lu. "CRISPR-Cas9 for in vivo gene ther- apy: Promise and hurdles." Molecular therapy Nucleic acids 5 (2016).
    View at Publisher | View at Google Scholar
19. Ding, Yuduan, Hong Li, Ling-Ling Chen, and Kabin Xie. "Re- cent advances in genome editing using CRISPR/Cas9." Fron- tiers in plant science 7 (2016): 703.
    View at Publisher | View at Google Scholar
20. Doench, John G., Nicolo Fusi, Meagan Sullender, Mudra Heg- de, Emma W. Vaimberg, Katherine F. Donovan, Ian Smith et al. "Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9." Nature biotechnology 34, no. 2 (2016): 184-191.
    View at Publisher | View at Google Scholar
21. Doman, Jordan L., Aditya Raguram, Gregory A. Newby, and David R. Liu. "Evaluation and minimization of Cas9-indepen- dent off-target DNA editing by cytosine base editors." Nature biotechnology 38, no. 5 (2020): 620-628.
    View at Publisher | View at Google Scholar
22. Donovan, Sophie, Yuwei Mao, Douglas J. Orr, Elizabete Car- mo-Silva, and Alistair J. McCormick. "CRISPR-Cas9-mediated mutagenesis of the Rubisco small subunit family in Nicotiana tabacum." Frontiers in Genome Editing 2 (2020): 605614.
    View at Publisher | View at Google Scholar
23. Doudna, Jennifer A., and Emmanuelle Charpentier. "The new frontier of genome engineering with CRISPR-Cas9." Science 346, no. 6213 (2014): 1258096.
    View at Publisher | View at Google Scholar
24. Durland, J., & Ahmadian-Moghadam, H. (2021). Genetics, mu- tagenesis. In StatPearls [Internet]. StatPearls Publishing.
    View at Publisher | View at Google Scholar
25. El-Gewely, M. Raafat, Chris Fenton, Elisabeth Kjeldsen, and Hao Xu. "Mutagenesis: site-specific." The Encyclopedia of Life Sciences (2001).
    View at Publisher | View at Google Scholar
26. Endo, Masaki, Ayako Nishizawa-Yokoi, and Seiichi Toki. "Rice genome editing." Rice genomics, genetics and breeding (2018): 523-539.
    View at Publisher | View at Google Scholar
27. Fang, Chuanying, Kang Li, Yangyang Wu, Dehong Wang, Jun- jie Zhou, Xiaoli Liu, Yufei Li et al. "OsTSD2-mediated cell wall modification affects ion homeostasis and salt tolerance." Plant, Cell & Environment 42, no. 5 (2019): 1503-1512.
    View at Publisher | View at Google Scholar
28. UNICEF. (2021). The state of food security and nutrition in the world 2021.
    View at Publisher | View at Google Scholar
29. Yunyan, Fei, Yang Jie, Wang Fangquan, Fan Fangjun, L. I. Wenqi, Wang Jun, X. U. Yang, Zhu Jinyan, and Zhong Weigong. "Production of two elite glutinous rice varieties by editing Wx gene." Rice Science 26, no. 2 (2019): 118-124.
    View at Publisher | View at Google Scholar
30. Fernie, Alisdair R., and Jianbing Yan. "De novo domestication: an alternative route toward new crops for the future." Molecular plant 12, no. 5 (2019): 615-631.
    View at Publisher | View at Google Scholar
31. Fiaz, Sajid, Xiukang Wang, Afifa Younas, Badr Alharthi, Adeel Riaz, and Habib Ali. "Apomixis and strategies to induce apo- mixis to preserve hybrid vigor for multiple generations." GM crops & food 12, no. 1 (2021): 57-70.
    View at Publisher | View at Google Scholar
32. Fineran, Peter C., and Emmanuelle Charpentier. "Memory of viral infections by CRISPR-Cas adaptive immune systems: ac- quisition of new information." Virology 434, no. 2 (2012): 202- 209.
    View at Publisher | View at Google Scholar
33. Fu, Yanfang, Jeffry D. Sander, Deepak Reyon, Vincent M. Cascio, and J. Keith Joung. "Improving CRISPR-Cas nuclease specificity using truncated guide RNAs." Nature biotechnology 32, no. 3 (2014): 279-284.
    View at Publisher | View at Google Scholar
34. Gaj, Thomas, Shannon J. Sirk, Sai-lan Shui, and Jia Liu. "Genome-editing technologies: principles and applications." Cold Spring Harbor perspectives in biology 8, no. 12 (2016): a023754.
    View at Publisher | View at Google Scholar
35. Gantz, Valentino M., and Ethan Bier. "The mutagenic chain re- action: a method for converting heterozygous to homozygous mutations." Science 348, no. 6233 (2015): 442-444.
    View at Publisher | View at Google Scholar
36. Gao, Linyi, David BT Cox, Winston X. Yan, John C. Mantei- ga, Martin W. Schneider, Takashi Yamano, Hiroshi Nishimasu, Osamu Nureki, Nicola Crosetto, and Feng Zhang. "Engineered Cpf1 variants with altered PAM specificities." Nature biotech- nology 35, no. 8 (2017): 789-792.
    View at Publisher | View at Google Scholar
37. MN, Bissah. "Medires Publishers-Article."
    View at Publisher | View at Google Scholar
38. Gomez, Michael A., Z. Daniel Lin, Theodore Moll, Raj Deepi- ka Chauhan, Luke Hayden, Kelley Renninger, Getu Beyene et al. "Simultaneous CRISPR/Cas9-mediated editing of cassava eIF 4E isoforms nCBP-1 and nCBP-2 reduces cassava brown streak disease symptom severity and incidence." Plant bio- technology journal 17, no. 2 (2019): 421-434.
    View at Publisher | View at Google Scholar
39. Haeussler, Maximilian, and Jean-Paul Concordet. "Genome editing with CRISPR-Cas9: can it get any better?." Journal of genetics and genomics 43, no. 5 (2016): 239-250.
    View at Publisher | View at Google Scholar
40. Halford, Nigel G. "Legislation governing genetically modified and genome-edited crops in Europe: the need for change." Journal of the Science of Food and Agriculture 99, no. 1 (2019): 8-12.
    View at Publisher | View at Google Scholar
41. Hand, Melanie L., and Anna MG Koltunow. "The genetic con- trol of apomixis: asexual seed formation." Genetics 197, no. 2 (2014): 441-450.
    View at Publisher | View at Google Scholar
42. Mahmudul, HassanMd, and A. TuskanGerald. "Prime editing technology and its prospects for future applications in plant bi- ology research." BioDesign Research (2020).
    View at Publisher | View at Google Scholar
43. Hojsgaard, Diego, and Elvira Hörandl. "A little bit of sex mat- ters for genome evolution in asexual plants." Frontiers in plant science 6 (2015): 82.
    View at Publisher | View at Google Scholar
44. Hua, Kai, Jinshan Zhang, Jose Ramon Botella, Changle Ma, Fanjiang Kong, Baohui Liu, and Jian-Kang Zhu. "Perspec- tives on the application of genome-editing technologies in crop breeding." Molecular Plant 12, no. 8 (2019): 1047-1059.
    View at Publisher | View at Google Scholar
45. Hua, Kai, Yuwei Jiang, Xiaoping Tao, and Jian-Kang Zhu. "Pre- cision genome engineering in rice using prime editing system." Plant Biotechnology Journal 18, no. 11 (2020): 2167.
    View at Publisher | View at Google Scholar
46. Huang, Teng-Kuei, and Holger Puchta. "CRISPR/Cas-medi- ated gene targeting in plants: finally a turn for the better for homologous recombination." Plant Cell Reports 38 (2019): 443-453.
    View at Publisher | View at Google Scholar
47. Jiang, Yu, Fenghui Qian, Junjie Yang, Yingmiao Liu, Feng Dong, Chongmao Xu, Bingbing Sun et al. "CRISPR-Cpf1 as- sisted genome editing of Corynebacterium glutamicum." Na- ture communications 8, no. 1 (2017): 15179.
    View at Publisher | View at Google Scholar
48. Jiang, Wenzhi, Huanbin Zhou, Honghao Bi, Michael Fromm, Bing Yang, and Donald P. Weeks. "Demonstration of CRISPR/ Cas9/sgRNA-mediated targeted gene modification in Arabi- dopsis, tobacco, sorghum and rice." Nucleic acids research 41, no. 20 (2013): e188-e188.
    View at Publisher | View at Google Scholar
49. Jiang, Yilin, Kangtai Sun, and Xueli An. "CRISPR/Cas system: applications and prospects for maize improvement." ACS Agri- cultural Science & Technology 2, no. 2 (2022): 174-183.
    View at Publisher | View at Google Scholar
50. Jinek, Martin, Krzysztof Chylinski, Ines Fonfara, Michael Hau- er, Jennifer A. Doudna, and Emmanuelle Charpentier. "A pro- grammable dual-RNA–guided DNA endonuclease in adaptive bacterial immunity." science 337, no. 6096 (2012): 816-821.
    View at Publisher | View at Google Scholar
51. Johnsson, Martin, R. Chris Gaynor, Janez Jenko, Gregor Gor- janc, Dirk-Jan De Koning, and John M. Hickey. "Removal of alleles by genome editing (RAGE) against deleterious load." Genetics Selection Evolution 51 (2019): 1-18.
    View at Publisher | View at Google Scholar
52. Jorasch, Petra. "Potential, challenges, and threats for the appli- cation of new breeding techniques by the private plant breeding sector in the EU." Frontiers in Plant Science 11 (2020): 582011.
    View at Publisher | View at Google Scholar
53. Kaushal, P., D. R. Malaviya, and A. K. Roy. "Prospects for breeding apomictic rice: a reassessment." Current Science 87, no. 3 (2004): 292-296.
    View at Publisher | View at Google Scholar
54. Khanday, Imtiyaz, Debra Skinner, Bing Yang, Raphael Merci- er, and Venkatesan Sundaresan. "A male-expressed rice em- bryogenic trigger redirected for asexual propagation through seeds." Nature 565, no. 7737 (2019): 91-95.
    View at Publisher | View at Google Scholar
55. Kim, Y. Bill, Alexis C. Komor, Jonathan M. Levy, Michael S. Packer, Kevin T. Zhao, and David R. Liu. "Increasing the ge- nome-targeting scope and precision of base editing with engi- neered Cas9-cytidine deaminase fusions." Nature biotechnolo- gy 35, no. 4 (2017): 371-376.
    View at Publisher | View at Google Scholar
56. Kleinstiver, Benjamin P., Michelle S. Prew, Shengdar Q. Tsai, Nhu T. Nguyen, Ved V. Topkar, Zongli Zheng, and J. Keith Joung. "Broadening the targeting range of Staphylococcus aureus CRISPR-Cas9 by modifying PAM recognition." Nature biotechnology 33, no. 12 (2015): 1293-1298.
    View at Publisher | View at Google Scholar
57. Koltunow, A. M., Ozias-Akins, P. and Siddiqi, I. (2013). Apo- mixis, pp. 83–110 in Seed Genomics, edited by P. W. Becraft. Wiley, New York
    View at Publisher | View at Google Scholar
58. Koltunow, Anna M., and Ueli Grossniklaus. "Apomixis: a devel- opmental perspective." Annual review of plant biology 54, no. 1 (2003): 547-574.
    View at Publisher | View at Google Scholar
59. Koonin, Eugene V., Kira S. Makarova, and Feng Zhang. "Di- versity, classification and evolution of CRISPR-Cas systems." Current opinion in microbiology 37 (2017): 67-78.
    View at Publisher | View at Google Scholar
60. Krenek, Pavel, Olga Samajova, Ivan Luptovciak, Anna Dosko- cilova, George Komis, and Jozef Samaj. "Transient plant trans- formation mediated by Agrobacterium tumefaciens: Principles, methods and applications." Biotechnology Advances 33, no. 6 (2015): 1024-1042.
    View at Publisher | View at Google Scholar
61. Kurata, Morito, Natalie K. Wolf, Walker S. Lahr, Madison T. Weg, Mitchell G. Kluesner, Samantha Lee, Kai Hui, Masano Shiraiwa, Beau R. Webber, and Branden S. Moriarity. "Highly multiplexed genome engineering using CRISPR/Cas9 gRNA arrays." PloS one 13, no. 9 (2018): e0198714.
    View at Publisher | View at Google Scholar
62. Lacchini, Elia, Edward Kiegle, Marco Castellani, Hélène Adam, Stefan Jouannic, Veronica Gregis, and Martin M. Kater. "CRIS- PR-mediated accelerated domestication of African rice landra- ces." PLoS One 15, no. 3 (2020): e0229782.
    View at Publisher | View at Google Scholar
63. Lawrenson, Tom, Oluwaseyi Shorinola, Nicola Stacey, Chen- gdao Li, Lars Østergaard, Nicola Patron, Cristobal Uauy, and Wendy Harwood. "Induction of targeted, heritable mutations in barley and Brassica oleracea using RNA-guided Cas9 nucle- ase." Genome biology 16 (2015): 1-13.
    View at Publisher | View at Google Scholar
64. Le, Ngoc Thu, Huyen Thi Tran, Thao Phuong Bui, Giang Thu Nguyen, Doai Van Nguyen, Dong Thi Ta, Duy Dinh Trinh et al. "Simultaneously induced mutations in eIF4E genes by CRIS- PR/Cas9 enhance PVY resistance in tobacco." Scientific Re- ports 12, no. 1 (2022): 14627.
    View at Publisher | View at Google Scholar
65. Lei, Yang, Li Lu, Hai-Yang Liu, Sen Li, Feng Xing, and Ling- Ling Chen. "CRISPR-P: a web tool for synthetic single-guide RNA design of CRISPR-system in plants." Molecular plant 7, no. 9 (2014): 1494-1496.
    View at Publisher | View at Google Scholar
66. Lei, Yang, Li Lu, Hai-Yang Liu, Sen Li, Feng Xing, and Ling- Ling Chen. "CRISPR-P: a web tool for synthetic single-guide RNA design of CRISPR-system in plants." Molecular plant 7, no. 9 (2014): 1494-1496.
    View at Publisher | View at Google Scholar
67. Li, Jun, Xiangbing Meng, Yuan Zong, Kunling Chen, Huawei Zhang, Jinxing Liu, Jiayang Li, and Caixia Gao. "Gene replace- ments and insertions in rice by intron targeting using CRISPR– Cas9." Nature plants 2, no. 10 (2016): 1-6.
    View at Publisher | View at Google Scholar
68. Li, Lei, Keke Wei, Guosong Zheng, Xiaocao Liu, Shaoxin Chen, Weihong Jiang, and Yinhua Lu. "CRISPR-Cpf1-assist- ed multiplex genome editing and transcriptional repression in Streptomyces." Applied and Environmental Microbiology 84, no. 18 (2018): e00827-18.
    View at Publisher | View at Google Scholar
69. Li, Jian-Feng, Julie E. Norville, John Aach, Matthew McCor- mack, Dandan Zhang, Jenifer Bush, George M. Church, and Jen Sheen. "Multiplex and homologous recombination–mediat- ed genome editing in Arabidopsis and Nicotiana benthamiana using guide RNA and Cas9." Nature biotechnology 31, no. 8 (2013): 688-691.
    View at Publisher | View at Google Scholar
70. Li, Jun, Yan Li, and Ligeng Ma. "Recent advances in CRIS- PR/Cas9 and applications for wheat functional genomics and breeding." Abiotech 2, no. 4 (2021): 375-385.
    View at Publisher | View at Google Scholar
71. Li, Shaoya, Xin Zhang, Wensheng Wang, Xiuping Guo, Zhichao Wu, Wenming Du, Yunde Zhao, and Lanqin Xia. "Expanding the scope of CRISPR/Cpf1-mediated genome editing in rice." Molecular Plant 11, no. 7 (2018): 995-998.
    View at Publisher | View at Google Scholar
72. Yu, Hong, Tao Lin, Xiangbing Meng, Huilong Du, Jingkun Zhang, Guifu Liu, Mingjiang Chen et al. "A route to de novo domestication of wild allotetraploid rice." Cell 184, no. 5 (2021): 1156-1170.
    View at Publisher | View at Google Scholar
73. Li, Jingying, Xin Zhang, Yongwei Sun, Jiahui Zhang, Wenming Du, Xiuping Guo, Shaoya Li, Yunde Zhao, and Lanqin Xia. "Ef- ficient allelic replacement in rice by gene editing: a case study of the NRT1. 1B gene." Journal of integrative plant biology 60, no. 7 (2018): 536-540.
    View at Publisher | View at Google Scholar
74. Li, Yuan, Zhen Lin, Yang Yue, Haiming Zhao, Xiaohong Fei, Lizhu E, Chenxu Liu, Shaojiang Chen, Jinsheng Lai, and Weib- in Song. "Loss-of-function alleles of ZmPLD3 cause haploid in- duction in maize." Nature plants 7, no. 12 (2021): 1579-1588.
    View at Publisher | View at Google Scholar
75. Li, Zhongsen, Zhan-Bin Liu, Aiqiu Xing, Bryan P. Moon, Jessi- ca P. Koellhoffer, Lingxia Huang, R. Timothy Ward, Elizabeth Clifton, S. Carl Falco, and A. Mark Cigan. "Cas9-guide RNA directed genome editing in soybean." Plant physiology 169, no. 2 (2015): 960-970.
    View at Publisher | View at Google Scholar
76. Liang, Zhen, Kunling Chen, and Caixia Gao. "Biolistic delivery of CRISPR/Cas9 with ribonucleoprotein complex in wheat." Plant genome editing with CRISPR systems: methods and protocols (2019): 327-335.
    View at Publisher | View at Google Scholar
77. Liang, Zhen, Kang Zhang, Kunling Chen, and Caixia Gao. "Tar- geted mutagenesis in Zea mays using TALENs and the CRIS- PR/Cas system." Journal of Genetics and Genomics 41, no. 2 (2014): 63-68.
    View at Publisher | View at Google Scholar
78. Liang, Zhen, Kunling Chen, Tingdong Li, Yi Zhang, Yanpeng Wang, Qian Zhao, Jinxing Liu et al. "Efficient DNA-free genome editing of bread wheat using CRISPR/Cas9 ribonucleoprotein complexes." Nature communications 8, no. 1 (2017): 14261.
    View at Publisher | View at Google Scholar
79. Lieber, Michael R. "The mechanism of double-strand DNA break repair by the nonhomologous DNA end-joining pathway." Annual review of biochemistry 79, no. 1 (2010): 181-211.
    View at Publisher | View at Google Scholar
80. Lin, Su-Ru, Hung-Chih Yang, Yi-Ting Kuo, Chun-Jen Liu, Ta- Yu Yang, Ku-Chun Sung, You-Yu Lin et al. "The CRISPR/Cas9 system facilitates clearance of the intrahepatic HBV templates in vivo." Molecular therapy Nucleic acids 3 (2014).
    View at Publisher | View at Google Scholar
81. Liu, Guoquan, Jieqing Li, and Ian D. Godwin. "Genome editing by CRISPR/Cas9 in sorghum through biolistic bombardment." Sorghum: Methods and protocols (2019): 169-183.
    View at Publisher | View at Google Scholar
82. Lowder, Levi G., Dengwei Zhang, Nicholas J. Baltes, Joseph W. Paul III, Xu Tang, Xuelian Zheng, Daniel F. Voytas, Tzu- ng-Fu Hsieh, Yong Zhang, and Yiping Qi. "A CRISPR/Cas9 toolbox for multiplexed plant genome editing and transcription- al regulation." Plant physiology 169, no. 2 (2015): 971-985.
    View at Publisher | View at Google Scholar
83. Lu, Yuming, Xiao Ye, Renming Guo, Jing Huang, Wei Wang, Jiuyou Tang, Longtao Tan, Jian-kang Zhu, Chengcai Chu, and Yangwen Qian. "Genome-wide targeted mutagenesis in rice using the CRISPR/Cas9 system." Molecular plant 10, no. 9 (2017): 1242-1245.
    View at Publisher | View at Google Scholar
84. Lucioli, Alessandra, Raffaela Tavazza, Simona Baima, Karoly Fatyol, Jozsef Burgyan, and Mario Tavazza. "CRISPR-Cas9 targeting of the eIF4E1 gene extends the potato virus y re- sistance spectrum of the Solanum tuberosum l. cv. desirée." Frontiers in Microbiology 13 (2022): 873930.
    View at Publisher | View at Google Scholar
85. Luo, Qiong, Yafei Li, Yi Shen, and Zhukuan Cheng. "Ten years of gene discovery for meiotic event control in rice." Journal of Genetics and Genomics 41, no. 3 (2014): 125-137.
    View at Publisher | View at Google Scholar
86. Lyons, Jessica B., Jessen V. Bredeson, Ben N. Mansfeld, Guil- laume Jean Bauchet, Jeffrey Berry, Adam Boyher, Lukas A. Mueller, Daniel S. Rokhsar, and Rebecca S. Bart. "Current sta- tus and impending progress for cassava structural genomics." Plant Molecular Biology (2021): 1-15.
    View at Publisher | View at Google Scholar
87. Ma, Xingliang, Qunyu Zhang, Qinlong Zhu, Wei Liu, Yan Chen, Rong Qiu, Bin Wang et al. "A robust CRISPR/Cas9 system for convenient, high-efficiency multiplex genome editing in mono- cot and dicot plants." Molecular plant 8, no. 8 (2015): 1274- 1284.
    View at Publisher | View at Google Scholar
88. Mao, Yanfei, Hui Zhang, Nanfei Xu, Botao Zhang, Feng Gou, and Jian-Kang Zhu. "Application of the CRISPR–Cas system for efficient genome engineering in plants." Molecular plant 6, no. 6 (2013): 2008-2011.
    View at Publisher | View at Google Scholar
89. Mao, Yanfei, Zhengjing Zhang, Zhengyan Feng, Pengliang Wei, Hui Zhang, José Ramón Botella, and Jian-Kang Zhu. "De- velopment of germ-line-specific CRISPR-Cas9 systems to im- prove the production of heritable gene modifications in Arabi- dopsis." Plant biotechnology journal 14, no. 2 (2016): 519-532
    View at Publisher | View at Google Scholar
90. Marimuthu, Mohan PA, Sylvie Jolivet, Maruthachalam Ravi, Lucie Pereira, Jayeshkumar N. Davda, Laurence Cromer, Lili Wang et al. "Synthetic clonal reproduction through seeds." Sci- ence 331, no. 6019 (2011): 876-876.
    View at Publisher | View at Google Scholar
91. Matsumoto, Takashi, Wu, Jianzhong, Kanamori, Hiroyuki, Ka- tayose, Yuichi,Fujisawa, Masaki, Namiki, Nobukazu, Mizuno, Hiroshi, Yamamoto, Kimiko, Antonio, Baltazar, Baba, Tomoya, Sakata, Katsumi, Nagamura, Yoshiaki, Aoki, Hiroyoshi, Arika- wa, Koji, Arita, KoheiBiti, Takahito, Chiden, Yoshino, Fujitsuka, Nahoko, Fukunaka, Rie and Burr, B. (2005). The map- based sequence of the rice genome. Nature 436:793-800
    View at Publisher | View at Google Scholar
92. Meng, Xiangbing, Hong Yu, Yi Zhang, Fengfeng Zhuang, Xiaoguang Song, Songsong Gao, Caixia Gao, and Jiayang Li. "Construction of a genome-wide mutant library in rice using CRISPR/Cas9." Molecular plant 10, no. 9 (2017): 1238-1241.
    View at Publisher | View at Google Scholar
93. Mercer, Andrew C., Thomas Gaj, Roberta P. Fuller, and Carlos F. Barbas III. "Chimeric TALE recombinases with programma- ble DNA sequence specificity." Nucleic acids research 40, no. 21 (2012): 11163-11172.
    View at Publisher | View at Google Scholar
94. Miao, Jin, Dongshu Guo, Jinzhe Zhang, Qingpei Huang, Genji Qin, Xin Zhang, Jianmin Wan, Hongya Gu, and Li-Jia Qu. "Tar- geted mutagenesis in rice using CRISPR-Cas system." Cell research 23, no. 10 (2013): 1233-1236.
    View at Publisher | View at Google Scholar
95. Mieulet, Delphine, Sylvie Jolivet, Maud Rivard, Laurence Cromer, Aurore Vernet, Pauline Mayonove, Lucie Pereira et al. "Turning rice meiosis into mitosis." Cell Research 26, no. 11 (2016): 1242-1254.
    View at Publisher | View at Google Scholar
96. Miller, Shannon M., Tina Wang, Peyton B. Randolph, Mandana Arbab, Max W. Shen, Tony P. Huang, Zaneta Matuszek, Greg- ory A. Newby, Holly A. Rees, and David R. Liu. "Continuous evolution of SpCas9 variants compatible with non-G PAMs." Nature biotechnology 38, no. 4 (2020): 471-481.
    View at Publisher | View at Google Scholar
97. Mishra, Rukmini, and Kaijun Zhao. "Genome editing technol- ogies and their applications in crop improvement." Plant Bio- technology Reports 12 (2018): 57-68.
    View at Publisher | View at Google Scholar
98. Mishra, Rukmini, Raj Kumar Joshi, and Kaijun Zhao. "Genome editing in rice: recent advances, challenges, and future implica- tions." Frontiers in Plant Science 9 (2018): 1361.
    View at Publisher | View at Google Scholar
99. Mohanta, Tapan Kumar, Tufail Bashir, Abeer Hashem, Elsayed Fathi Abd_Allah, and Hanhong Bae. "Genome editing tools in plants." Genes 8, no. 12 (2017): 399.
    View at Publisher | View at Google Scholar
100. Murovec, Jana, Katja Guček, Borut Bohanec, Monika Avbelj, and Roman Jerala. "DNA-free genome editing of Brassica oler- acea and B. rapa protoplasts using CRISPR-Cas9 ribonucleo- protein complexes." Frontiers in Plant Science 9 (2018): 1594.
     View at Publisher | View at Google Scholar
101. Odipio, John, Titus Alicai, Ivan Ingelbrecht, Dmitri A. Nusinow, Rebecca Bart, and Nigel J. Taylor. "Efficient CRISPR/Cas9 ge- nome editing of phytoene desaturase in cassava." Frontiers in plant science 8 (2017): 1780.
     View at Publisher | View at Google Scholar
102. Ogaugwu, Christian E., Stanley O. Agbo, and Modinat A. Ade- koya. "CRISPR in sub-Saharan Africa: applications and educa- tion." Trends in biotechnology 37, no. 3 (2019): 234-237.
     View at Publisher | View at Google Scholar
103. MN, Bissah. "Medires Publishers-Article."
     View at Publisher | View at Google Scholar
104. Pickar-Oliver, Adrian, and Charles A. Gersbach. "The next gen- eration of CRISPR–Cas technologies and applications." Na- ture reviews Molecular cell biology 20, no. 8 (2019): 490-507.
     View at Publisher | View at Google Scholar
105. Ravi, Maruthachalam, and Simon WL Chan. "Haploid plants produced by centromere-mediated genome elimination." Na- ture 464, no. 7288 (2010): 615-618.
     View at Publisher | View at Google Scholar
106. Ravi, Maruthachalam, Mohan PA Marimuthu, and Imran Sid- diqi. "Gamete formation without meiosis in Arabidopsis." Na- ture 451, no. 7182 (2008): 1121-1124.
     View at Publisher | View at Google Scholar
107. Romero, Fernando Matías, and Andrés Gatica-Arias. "CRIS- PR/Cas9: development and application in rice breeding." Rice Science 26, no. 5 (2019): 265-281.
     View at Publisher | View at Google Scholar
108. Chow, R. D., Chen, J. S., Shen, J., & Chen, S. (2020). peg- Finder: A pegRNA designer for CRISPR prime editing. BioRxiv, 2020-05.
     View at Publisher | View at Google Scholar
109. Sailer, Christian, Bernhard Schmid, and Ueli Grossniklaus. "Apomixis allows the transgenerational fixation of phenotypes in hybrid plants." Current biology 26, no. 3 (2016): 331-337.
     View at Publisher | View at Google Scholar
110. San Filippo, Joseph, Patrick Sung, and Hannah Klein. "Mech- anism of eukaryotic homologous recombination." Annu. Rev. Biochem. 77, no. 1 (2008): 229-257.
     View at Publisher | View at Google Scholar
111. Sathee, Lekshmy, B. Jagadhesan, Pratheek H. Pandesha, Di- pankar Barman, Sandeep Adavi B, Shivani Nagar, G. K. Krish- na, Shailesh Tripathi, Shailendra K. Jha, and Viswanathan Chinnusamy. "Genome editing targets for improving nutrient use efficiency and nutrient stress adaptation." Frontiers in Ge- netics 13 (2022): 900897.
     View at Publisher | View at Google Scholar
112. Schulman, Alan H., Kirsi-Marja Oksman-Caldentey, and Teemu H. Teeri. "European Court of Justice delivers no justice to Eu- rope on genome-edited crops." Plant biotechnology journal 18, no. 1 (2019): 8.
     View at Publisher | View at Google Scholar
113. Sedlar, Aleš, Mateja Zupin, Marko Maras, Jaka Razinger, Jelka Šuštar-Vozlič, Barbara Pipan, and Vladimir Meglič. "QTL map- ping for drought-responsive agronomic traits associated with physiology, phenology, and yield in an Andean intra-gene pool common bean population." Agronomy 10, no. 2 (2020): 225.
     View at Publisher | View at Google Scholar
114. Sharma, Rita, Yan Liang, Mi Yeon Lee, Venkataramana R. Pi- datala, Jenny C. Mortimer, and Henrik V. Scheller. "Agrobacte- rium-mediated transient transformation of sorghum leaves for accelerating functional genomics and genome editing studies." BMC research notes 13 (2020): 1-7.
     View at Publisher | View at Google Scholar
115. Shibata, Atsushi. "Regulation of repair pathway choice at two-ended DNA double-strand breaks." Mutation Research/ Fundamental and Molecular Mechanisms of Mutagenesis 803 (2017): 51-55.
     View at Publisher | View at Google Scholar
116. Shmakov, Sergey, Aaron Smargon, David Scott, David Cox, Neena Pyzocha, Winston Yan, Omar O. Abudayyeh et al. "Di- versity and evolution of class 2 CRISPR–Cas systems." Nature reviews microbiology 15, no. 3 (2017): 169-182.
     View at Publisher | View at Google Scholar
117. Sikora, P., Chawade, A., Larsson, M., Olsson, J., & Olsson, O. (2011). Mutagenesis as a tool in plant genetics, functional ge- nomics, and breeding. International journal of plant genomics, 2011.
     View at Publisher | View at Google Scholar
118. Sun, Yongwei, Guiai Jiao, Zupei Liu, Xin Zhang, Jingying Li, Xiuping Guo, Wenming Du et al. "Generation of high-amylose rice through CRISPR/Cas9-mediated targeted mutagenesis of starch branching enzymes." Frontiers in plant science 8 (2017): 298.
     View at Publisher | View at Google Scholar
119. Sun, Yongwei, Guiai Jiao, Zupei Liu, Xin Zhang, Jingying Li, Xiuping Guo, Wenming Du et al. "Generation of high-amylose rice through CRISPR/Cas9-mediated targeted mutagenesis of starch branching enzymes." Frontiers in plant science 8 (2017): 298.
     View at Publisher | View at Google Scholar
120. Sun, Yongwei, Guiai Jiao, Zupei Liu, Xin Zhang, Jingying Li, Xiuping Guo, Wenming Du et al. "Generation of high-amylose rice through CRISPR/Cas9-mediated targeted mutagenesis of starch branching enzymes." Frontiers in plant science 8 (2017): 298.
     View at Publisher | View at Google Scholar
121. Svitashev, Sergei, Joshua K. Young, Christine Schwartz, Huirong Gao, S. Carl Falco, and A. Mark Cigan. "Targeted mu- tagenesis, precise gene editing, and site-specific gene inser- tion in maize using Cas9 and guide RNA." Plant physiology 169, no. 2 (2015): 931-945.
     View at Publisher | View at Google Scholar
122. Andoh, Cletus Tandoh. "Genome editing technologies: ethical and regulation challenges for Africa." International Journal of Health Economics and Policy 2, no. 2 (2017): 30-46.
     View at Publisher | View at Google Scholar
123. Tang, Tian, Yingli Han, Yuran Wang, He Huang, and Pengxu Qian. "Programmable system of Cas13-mediated RNA modifi- cation and its biological and biomedical applications." Frontiers in Cell and Developmental Biology 9 (2021): 677587.
     View at Publisher | View at Google Scholar
124. Thiruppathi, Dhineshkumar. "Maize RNA polymerase III sub- unit NRPC2: new kid on the kernel development block." (2020): 12-13.
     View at Publisher | View at Google Scholar
125. Tian, Yinshuai, Xinanbei Liu, Caixin Fan, Tingting Li, Huan Qin, Xiao Li, Kai Chen, Yunpu Zheng, Fang Chen, and Ying Xu. "Enhancement of tobacco (Nicotiana tabacum L.) seed lipid content for biodiesel production by CRISPR-Cas9-medi- ated knockout of NtAn1." Frontiers in Plant Science 11 (2021): 599474.
     View at Publisher | View at Google Scholar
126. Tian, Yinshuai, Kai Chen, Xiao Li, Yunpu Zheng, and Fang Chen. "Design of high-oleic tobacco (Nicotiana tabacum L.) seed oil by CRISPR-Cas9-mediated knockout of NtFAD2–2." BMC Plant Biology 20 (2020): 1-12.
     View at Publisher | View at Google Scholar
127. Schardl, Christopher L., Simona Florea, Juan Pan, Padmaja Nagabhyru, Sladana Bec, and Patrick J. Calie. "The epichloae: alkaloid diversity and roles in symbiosis with grasses." Current opinion in plant biology 16, no. 4 (2013): 480-488.
     View at Publisher | View at Google Scholar
128. Tripathi, Leena, Valentine O. Ntui, and Jaindra N. Tripathi. "CRISPR/Cas9-based genome editing of banana for disease resistance." Current Opinion in Plant Biology 56 (2020): 118- 126.
     View at Publisher | View at Google Scholar
129. Tripathi, Leena, John Odipio, Jaindra Nath Tripathi, and Geof- frey Tusiime. "A rapid technique for screening banana cultivars for resistance to Xanthomonas wilt." European Journal of Plant Pathology 121 (2008): 9-19.
     View at Publisher | View at Google Scholar
130. Tripathi, Leena, Valentine Otang Ntui, Jaindra Nath Tripathi, and P. Lava Kumar. "Application of CRISPR/Cas for diagnosis and management of viral diseases of banana." Frontiers in Mi- crobiology 11 (2021): 609784.
     View at Publisher | View at Google Scholar
131. Yuan, Yuan, Wei Wen, Susan E. Yost, Quanhua Xing, Jin Yan, Ernest S. Han, Joanne Mortimer, and John H. Yim. "Combi- nation therapy with BYL719 and LEE011 is synergistic and causes a greater suppression of p-S6 in triple negative breast cancer." Scientific reports 9, no. 1 (2019): 7509.
     View at Publisher | View at Google Scholar
132. Tripathi, Leena, Valentine O. Ntui, and Jaindra N. Tripathi. "Control of bacterial diseases of banana using CRISPR/Cas- based gene editing." International Journal of Molecular Scienc- es 23, no. 7 (2022): 3619.
     View at Publisher | View at Google Scholar
133. Veillet, Florian, Laura Perrot, Laura Chauvin, Marie-Paule Ker- marrec, Anouchka Guyon-Debast, Jean-Eric Chauvin, Fabien Nogué, and Marianne Mazier. "Transgene-free genome edit- ing in tomato and potato plants using agrobacterium-mediated delivery of a CRISPR/Cas9 cytidine base editor." International Journal of Molecular Sciences 20, no. 2 (2019): 402.
     View at Publisher | View at Google Scholar
134. Veillet, Florian, Marie-Paule Kermarrec, Laura Chauvin, Jean-Eric Chauvin, and Fabien Nogué. "CRISPR-induced in- dels and base editing using the Staphylococcus aureus Cas9 in potato." PLoS One 15, no. 8 (2020): e0235942.
     View at Publisher | View at Google Scholar
135. Voytas, Daniel F. "Plant genome engineering with se- quence-specific nucleases." Annual review of plant biology 64, no. 1 (2013): 327-350.
     View at Publisher | View at Google Scholar
136. Voytas, Daniel F., and Caixia Gao. "Precision genome engi- neering and agriculture: opportunities and regulatory challeng- es." PLoS biology 12, no. 6 (2014): e1001877.
     View at Publisher | View at Google Scholar
137. Wada, Naoki, Risa Ueta, Yuriko Osakabe, and Keishi Osak- abe. "Precision genome editing in plants: state-of-the-art in CRISPR/Cas9-based genome engineering." BMC Plant Biol- ogy 20 (2020): 1-12.
     View at Publisher | View at Google Scholar
138. Wang, Fujun, Chunlian Wang, Piqing Liu, Cailin Lei, Wei Hao, Ying Gao, Yao-Guang Liu, and Kaijun Zhao. "Enhanced rice blast resistance by CRISPR/Cas9-targeted mutagenesis of the ERF transcription factor gene OsERF922." PloS one 11, no. 4 (2016): e0154027.
     View at Publisher | View at Google Scholar
139. Wang, Fujun, Chunlian Wang, Piqing Liu, Cailin Lei, Wei Hao, Ying Gao, Yao-Guang Liu, and Kaijun Zhao. "Enhanced rice blast resistance by CRISPR/Cas9-targeted mutagenesis of the ERF transcription factor gene OsERF922." PloS one 11, no. 4 (2016): e0154027.
     View at Publisher | View at Google Scholar
140. Wang, Hai, Shijuan Yan, Hongjia Xin, Wenjie Huang, Hao Zhang, Shouzhen Teng, Ya-Chi Yu et al. "A subsidiary cell-lo- calized glucose transporter promotes stomatal conductance and photosynthesis." The Plant Cell 31, no. 6 (2019): 1328- 1343.
     View at Publisher | View at Google Scholar
141. Wang, Xia, Yuantao Xu, Siqi Zhang, Li Cao, Yue Huang, Jun- feng Cheng, Guizhi Wu et al. "Genomic analyses of primitive, wild and cultivated citrus provide insights into asexual repro- duction." Nature genetics 49, no. 5 (2017): 765-772.
     View at Publisher | View at Google Scholar
142. Wang, Ying, Lizhao Geng, Menglong Yuan, Juan Wei, Chen Jin, Min Li, Kun Yu et al. "Deletion of a target gene in Indica rice via CRISPR/Cas9." Plant Cell Reports 36 (2017): 1333-1343.
     View at Publisher | View at Google Scholar
143. Wang, Chun, Qing Liu, Yi Shen, Yufeng Hua, Junjie Wang, Ji- anrong Lin, Mingguo Wu et al. "Clonal seeds from hybrid rice by simultaneous genome engineering of meiosis and fertiliza- tion genes." Nature biotechnology 37, no. 3 (2019): 283-286.
     View at Publisher | View at Google Scholar
144. Wang, Kejian. "Fixation of hybrid vigor in rice: synthetic apo- mixis generated by genome editing." Abiotech 1, no. 1 (2020): 15-20.
     View at Publisher | View at Google Scholar
145. Wang, Meng, Shubin Wang, Zhen Liang, Weiming Shi, Caixia Gao, and Guangmin Xia. "From genetic stock to genome ed- iting: gene exploitation in wheat." Trends in Biotechnology 36, no. 2 (2018): 160-172.
     View at Publisher | View at Google Scholar
146. Wang, Yanpeng, Xi Cheng, Qiwei Shan, Yi Zhang, Jinxing Liu, Caixia Gao, and Jin-Long Qiu. "Simultaneous editing of three homoeoalleles in hexaploid bread wheat confers heritable re- sistance to powdery mildew." Nature biotechnology 32, no. 9 (2014): 947-951.
     View at Publisher | View at Google Scholar
147. Wolt, Jeffrey D., Kan Wang, and Bing Yang. "The regulatory status of genome-edited crops." Plant biotechnology journal 14, no. 2 (2016): 510-518.
     View at Publisher | View at Google Scholar
148. Woo, Je Wook, Jungeun Kim, Soon Il Kwon, Claudia Corvalán, Seung Woo Cho, Hyeran Kim, Sang-Gyu Kim, Sang-Tae Kim, Sunghwa Choe, and Jin-Soo Kim. "DNA-free genome editing in plants with preassembled CRISPR-Cas9 ribonucleopro- teins." Nature biotechnology 33, no. 11 (2015): 1162-1164.
     View at Publisher | View at Google Scholar
149. Wu, Shao-Shuai, Qing-Cui Li, Chang-Qing Yin, Wen Xue, and Chun-Qing Song. "Advances in CRISPR/Cas-based gene therapy in human genetic diseases." Theranostics 10, no. 10 (2020): 4374.
     View at Publisher | View at Google Scholar
150. Wu, Shao-Shuai, Qing-Cui Li, Chang-Qing Yin, Wen Xue, and Chun-Qing Song. "Advances in CRISPR/Cas-based gene therapy in human genetic diseases." Theranostics 10, no. 10 (2020): 4374.
     View at Publisher | View at Google Scholar
151. Xie, En, Yafei Li, Ding Tang, Yanli Lv, Yi Shen, and Zhukuan Cheng. "A strategy for generating rice apomixis by gene edit- ing." Journal of integrative plant biology 61, no. 8 (2019): 911- 916.
     View at Publisher | View at Google Scholar
152. Xie, Ke, Suowei Wu, Ziwen Li, Yan Zhou, Danfeng Zhang, Zhenying Dong, Xueli An et al. "Map-based cloning and char- acterization of Zea mays male sterility33 (ZmMs33) gene, en- coding a glycerol-3-phosphate acyltransferase." Theoretical and Applied Genetics 131 (2018): 1363-1378.
     View at Publisher | View at Google Scholar
153. Yang, Chao, Yuki Hamamura, Kostika Sofroni, Franziska Böw- er, Sara Christina Stolze, Hirofumi Nakagami, and Arp Schnitt- ger. "SWITCH 1/DYAD is a WINGS APART-LIKE antagonist that maintains sister chromatid cohesion in meiosis." Nature communications 10, no. 1 (2019): 1755.
     View at Publisher | View at Google Scholar
154. Yang, Tianxia, Muhammad Ali, Lihao Lin, Ping Li, Hongju He, Qiang Zhu, Chuanlong Sun et al. "Recoloring tomato fruit by CRISPR/Cas9-mediated multiplex gene editing." Horticulture Research 10, no. 1 (2023): uhac214.
     View at Publisher | View at Google Scholar
155. Yang, Tian-xia, Lei Deng, Wei Zhao, Ruoxi Zhang, Hong-ling Jiang, Zhi-biao Ye, Chang-Bao Li, and Chuan-you Li. "Rapid breeding of pink-fruited tomato hybrids using the CRISPR/ Cas9 system." (2019): 505-508.
     View at Publisher | View at Google Scholar
156. Yao, Li, Ya Zhang, Chunxia Liu, Yubo Liu, Yanli Wang, Dawei Liang, Juntao Liu, Gayatri Sahoo, and Timothy Kelliher. "Os- MATL mutation induces haploid seed formation in indica rice." Nature plants 4, no. 8 (2018): 530-533.
     View at Publisher | View at Google Scholar
157. Yin, Pei Pei, Li Ping Tang, Xian Sheng Zhang, and Ying Hua Su. "Options for engineering apomixis in plants." Frontiers in Plant Science 13 (2022): 864987.
     View at Publisher | View at Google Scholar
158. Yuan, Guoqiang, Ting Zou, Xu Zhang, Miaomiao Liu, Tingting Luo, Zhiyuan He, Yang Tao et al. "A rice GDSL esterase/lipase protein (GELP) is required for anther and pollen development." Molecular Breeding 40 (2020): 1-15.
     View at Publisher | View at Google Scholar
159. Zhang, Anning, Yi Liu, Feiming Wang, Tianfei Li, Zhihao Chen, Deyan Kong, Junguo Bi et al. "Enhanced rice salinity toler- ance via CRISPR/Cas9-targeted mutagenesis of the OsRR22 gene." Molecular breeding 39 (2019): 1-10.
     View at Publisher | View at Google Scholar
160. Zhang, Jinshan, Hui Zhang, José Ramón Botella, and Ji- an-Kang Zhu. "Generation of new glutinous rice by CRISPR/ Cas9-targeted mutagenesis of the Waxy gene in elite rice va- rieties." Journal of integrative plant biology 60, no. 5 (2018): 369-375.
     View at Publisher | View at Google Scholar
161. Zhang, Jian-Hua, Poorni Adikaram, Mritunjay Pandey, Allison Genis, and William F. Simonds. "Optimization of genome edit- ing through CRISPR-Cas9 engineering." Bioengineered 7, no. 3 (2016): 166-174.
     View at Publisher | View at Google Scholar
162. Zhang, Anning, Yi Liu, Feiming Wang, Tianfei Li, Zhihao Chen, Deyan Kong, Junguo Bi et al. "Enhanced rice salinity toler- ance via CRISPR/Cas9-targeted mutagenesis of the OsRR22 gene." Molecular breeding 39 (2019): 1-10.
     View at Publisher | View at Google Scholar
163. Montagnani, C., R. Gentili, Matt Smith, M. F. Guarino, and S. Citterio. "The worldwide spread, success, and impact of rag- weed (Ambrosia spp.)." Critical Reviews in Plant Sciences 36, no. 3 (2017): 139-178.
     View at Publisher | View at Google Scholar
164. Zhang, Peipei, Yingxin Zhang, Lianping Sun, Sittipun Sinum- porn, Zhengfu Yang, Bin Sun, Dandan Xuan et al. "The rice AAA-ATPase OsFIGNL1 is essential for male meiosis." Fron- tiers in Plant Science 8 (2017): 1639.
     View at Publisher | View at Google Scholar
165. Zhang, Yi, Mathias Pribil, Michael Palmgren, and Caixia Gao. "A CRISPR way for accelerating improvement of food crops." Nature Food 1, no. 4 (2020): 200-205.
     View at Publisher | View at Google Scholar
166. Zhao, Hailiang, Yao Qin, Ziyi Xiao, Qi Li, Ning Yang, Zhenyuan Pan, Dianming Gong et al. "Loss of function of an RNA poly- merase III subunit leads to impaired maize kernel develop- ment." Plant physiology 184, no. 1 (2020): 359-373.
     View at Publisher | View at Google Scholar
167. Chen, Longzheng, Wei Li, Lorenzo Katin-Grazzini, Jing Ding, Xianbin Gu, Yanjun Li, Tingting Gu et al. "A method for the pro- duction and expedient screening of CRISPR/Cas9-mediated non-transgenic mutant plants." Horticulture research 5 (2018).
     View at Publisher | View at Google Scholar
168. Zheng, Shaoyan, Chanjuan Ye, Jingqin Lu, Jiamin Liufu, Lin Lin, Zequn Dong, Jing Li, and Chuxiong Zhuang. "Improving the rice photosynthetic efficiency and yield by editing OsHXK1 via CRISPR/Cas9 system." International journal of molecular sciences 22, no. 17 (2021): 9554.
     View at Publisher | View at Google Scholar
169. Zhong, Xuanbo, Wei Hong, Yue Shu, Jianfei Li, Lulu Liu, Xi- aoyang Chen, Faisal Islam, Weijun Zhou, and Guixiang Tang. "CRISPR/Cas9 mediated gene-editing of GmHdz4 transcrip- tion factor enhances drought tolerance in soybean (Glycine max [L.] Merr.)." Frontiers in Plant Science 13 (2022): 988505.
     View at Publisher | View at Google Scholar
170. Zhong, Yu, Chenxu Liu, Xiaolong Qi, Yanyan Jiao, Dong Wang, Yuwen Wang, Zongkai Liu et al. "Mutation of ZmDMP enhanc- es haploid induction in maize." Nature plants 5, no. 6 (2019): 575-580.
     View at Publisher | View at Google Scholar
171. Zhou, Xiaogang, Haicheng Liao, Mawsheng Chern, Junjie Yin, Yufei Chen, Jianping Wang, Xiaobo Zhu et al. "Loss of function of a rice TPR-domain RNA-binding protein confers broad-spec- trum disease resistance." Proceedings of the National Acade- my of Sciences 115, no. 12 (2018): 3174-3179.
     View at Publisher | View at Google Scholar
172. Zong, Yuan, Yijing Liu, Chenxiao Xue, Boshu Li, Xiangyang Li, Yanpeng Wang, Ji Li et al. "An engineered prime editor with enhanced editing efficiency in plants." Nature Biotechnology 40, no. 9 (2022): 1394-1402.
     View at Publisher | View at Google Scholar
173. Zong, Yuan, Yanpeng Wang, Chao Li, Rui Zhang, Kunling Chen, Yidong Ran, Jin-Long Qiu, Daowen Wang, and Caix- ia Gao. "Precise base editing in rice, wheat and maize with a Cas9-cytidine deaminase fusion." Nature biotechnology 35, no. 5 (2017): 438-440.
     View at Publisher | View at Google Scholar