Genome Engineering for Crop Improvement. Группа авторов

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СКАЧАТЬ Virus resistance Chandrasekaran et al. (2016) Camelina sativa FAD2 Low polyunsaturated fatty acids Jiang et al. (2017) Linumusitatissimum EPSPS Herbicide resistance Sauer et al. (2016) Carrot DcPDS and DcMYB113‐like genes depigmented carrot plants Xu et al. (2019) Strawberry PDS Carotenoid Biosynthesis Wilson et al. (2019) Grapes VvPDS Carotenoid Biosynthesis Nakajima et al. (2017) CsLOB1 Increased resistance to citrus canker Jia et al. (2017) MLO‐1 Negative regulator of resistance to Powdery mildew Malnoy et al. (2016) Pear MdPDS and TFL1.1 Carotenoid Biosynthesis & Floral repression Charrier et al. (2019) Apple PDS Carotenoid Biosynthesis Charrier et al. (2019, Nishitani et al. (2016) DIMP‐1/2/3 Negative regulator of resistance to blight disease Malnoy et al. (2016) Banana eBSV Resistance against Banana streak virus (eBSV) Tripathi et al. (2019) Chicory phytoene desaturase gene (CiPDS) Fruit ripening Jansing et al. (2019) Rubber HbFT HbTFL1 Delayed‐flowering and early‐flowering Fan et al. (2020) Brassica napus L.) BnLPAT2 and BnLPAT5 size of the oil bodies increased Zhang et al. (2019) Populus phytoene desaturase gene 8 Carotenoid Biosynthesis Fan et al. (2015) Flax EPSPS Aromatic amino acid biosynthesis Sauer et al. (2016)

1.5 CRISPR‐Cpf1

Introduction of CRISPR‐Cpf1 also known as CRISPR‐Cas12a further diversified the genome engineering methods (Figure 1.2). Cpf1 is an endonuclease which belongs to the class II CRISPR family (Alok et al. 2020; Zaidi et al. 2017). It was identified from Prevotella and Francisella1, therefore named as named as Cpf1. This system become popular as it was able to fill the gaps of previous genome editing tools and can substitute the CRISPR‐Cas9 in an efficient way (Moon et al. 2018). It is smaller in size than Cas9 and required a shorter CRISPR RNA for proper functioning (Liu et al. 2017). Unlike the CRISPR/Cas9, tracrRNA is no longer necessary to process Cpf1 associated mature CRISPR RNAs (Zetsche et al. 2015). Further, in contrast to the G rich PAM at 3′ end in CRISPR Cas9, it requires a T‐rich PAM sequence at the 5′‐end to perform cleavage efficiently, which enables the targeting of AT‐rich regions in the genome. Further, Cpf1 creates staggered cuts where insertion of a DNA fragment is easily possible via HDR (Gao et al. 2017). The off‐target binding of CRIPSR‐Cpf1 is comparatively less than CRISPR‐Cas9 which could be an additional benefit (Kim et al. 2016; Kleinstiver et al. 2016; Yan et al. 2017). A number of studies showed the application of CRISPR‐Cpf1 for targeted genome editing in various eukaryotes including plants (Kim et al. 2017; Zetsche et al. 2015). It has also been used for multiple targets in the genome (Wang et al. 2017).

Figure 1.2 Comparison of various features of CRISPR‐Cpf1 (a) and CRISPR‐Cas9.

      Source: Adapted from Zaidi et al. (2017) © 2017. Reproduced with the permission of Elsevier.

1.6 Conclusions

Genome‐editing technologies enable us to make precise changes in the genome of any living organism. These changes may be diverse‐insertion, deletion or even replacement of a particular stretch of DNA from the genome. Targeting these changes in accordance with our needs has always been the ambition of the scientific community. It was a challenging job until the development of recent genome‐editing technologies. Among these, the most popular and successful ones till now have been ZFNs, TALENs, and CRISPR‐Cas9. The breakthrough in the field of genome editing came after the discovery of CRISPR/Cas9 system as it is an RNA‐ guided and easy‐to‐design system. This system is fascinatingly repurposed as a genome editing tool and is till now the most efficient, cost effective and least demanding genome editing technique. Moreover, the recent addition of CRISPR‐Cpf1 that is a variant of the CRISPR‐Cas system further diversified the application of genome engineering tools by overcoming the various shortcomings of earlier systems.

Acknowledgements

      Ms. Sushmita is grateful to DST‐INSPIRE fellowship Program, DST, New Delhi, India for providing financial support. Authors are also thankful to CSIR, New Delhi for financial support in the form of “FBR Genome Editing Network Project” (MLP‐007).

      Institute's Manuscript Number is 'CSIR‐NBRI_MS/2020/06/24.

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