Improved Base Editor for Efficiently Inducing Genetic Variations in Rice with CRISPR/Cas9-Guided Hyperactive hAID Mutant

Ren, Bin; Yan, Fang; Kuang, Yongjie; Li, Na; Zhang, Dawei; Zhou, Xueping; Lin, Honghui

doi:10.1016/j.molp.2018.01.005

Cited by 181 publications

(104 citation statements)

References 10 publications

(20 reference statements)

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“…The efficiency of base-editing is enhanced using a nickase instead of catalytically inactive Cas9 that nicks the unmodified strand, thus stimulating mismatch repair using the unmodified strand as template [45][46][47][48]. The latest genome editing approach, however, enables the programmed conversion of single bases without the induction of DSBs by targeting deaminases to the site of interest.…”

Section: Using Cas9 As a Nickasementioning

confidence: 99%

“…By either fusing a cytidine deaminase or an adenosine deaminase to the Cas9 nickase a conversion of C/G to T/A or A/T to G/C could be induced. The efficiency of base-editing is enhanced using a nickase instead of catalytically inactive Cas9 that nicks the unmodified strand, thus stimulating mismatch repair using the unmodified strand as template [45][46][47][48]. Just recently, in addition to cytidine deaminases, adenosine deaminases could also be successfully employed for base-editing in plants [49].…”

Section: Using Cas9 As a Nickasementioning

confidence: 99%

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Transforming plant biology and breeding with CRISPR/Cas9, Cas12 and Cas13

2018

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Currently, biology is revolutionized by ever growing applications of the CRISPR/Cas system. As discussed in this Review, new avenues have opened up for plant research and breeding by the use of the sequence-specific DNases Cas9 and Cas12 (formerly named Cpf1) and, more recently, the RNase Cas13 (formerly named C2c2). Although double strand break-induced gene editing based on error-prone nonhomologous end joining has been applied to obtain new traits, such as powdery mildew resistance in wheat or improved pathogen resistance and increased yield in tomato, improved technologies based on CRISPR/Cas for programmed change in plant genomes via homologous recombination have recently been developed. Cas9- and Cas12- mediated DNA binding is used to develop tools for many useful applications, such as transcriptional regulation or fluorescence-based imaging of specific chromosomal loci in plant genomes. Cas13 has recently been applied to degrade mRNAs and combat viral RNA replication. By the possibility to address multiple sequences with different guide RNAs and by the simultaneous use of different Cas proteins in a single cell, we should soon be able to achieve complex changes of plant metabolism in a controlled way.

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Section: Using Cas9 As a Nickasementioning

confidence: 99%

Section: Using Cas9 As a Nickasementioning

confidence: 99%

Transforming plant biology and breeding with CRISPR/Cas9, Cas12 and Cas13

2018

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“…In some cases, homology directed repair (HDR) was used for targeted gene replacement (Endo et al ., 2016b; Gil‐Humanes et al ., ; Li et al ., ; Miki et al ., ; Schiml et al ., ; Svitashev et al ., ). Cas9 nickase (Fauser et al ., ; Ran et al ., ) and base editors (Gaudelli et al ., ; Komor et al ., ; Li et al ., ; Shimatani et al ., ) have further expanded the applications of Cas9‐based plant genome editing (Hua et al ., ; Lowder et al ., ; Lu and Zhu, ; Ren et al ., , ; Yan et al ., ; Zong et al ., ).…”

Section: Introductionmentioning

confidence: 99%

Single transcript unit CRISPR 2.0 systems for robust Cas9 and Cas12a mediated plant genome editing

Tang

Ren

Yang

et al. 2019

Plant Biotechnology Journal

127

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Summary CRISPR ‐Cas9 and Cas12a are two powerful genome editing systems. Expression of CRISPR in plants is typically achieved with a mixed dual promoter system, in which Cas protein is expressed by a Pol II promoter and a guide RNA is expressed by a species‐specific Pol III promoter such as U6 or U3. To achieve coordinated expression and compact vector packaging, it is desirable to express both CRISPR components under a single Pol II promoter. Previously, we demonstrated a first‐generation single transcript unit ( STU )‐Cas9 system, STU ‐Cas9‐ RZ , which is based on hammerhead ribozyme for processing single guide RNA s (sg RNA s). In this study, we developed two new STU ‐Cas9 systems and one STU ‐Cas12a system for applications in plants, collectively called the STU CRISPR 2.0 systems. We demonstrated these systems for genome editing in rice with both transient expression and stable transgenesis. The two STU ‐Cas9 2.0 systems process the sg RNA s with Csy4 ribonuclease and endogenous tRNA processing system respectively. Both STU ‐Cas9‐Csy4 and STU ‐Cas9‐ tRNA systems showed more robust genome editing efficiencies than our first‐generation STU ‐Cas9‐ RZ system and the conventional mixed dual promoter system. We further applied the STU ‐Cas9‐ tRNA system to compare two C to T base editing systems based on rAPOBEC 1 and Pm CDA 1 cytidine deaminases. The results suggest STU ‐based Pm CDA 1 base editor system is highly efficient in rice. The STU ‐Cas12a system, based on Cas12a’ self‐processing of a CRISPR RNA (cr RNA ) array, was also developed and demonstrated for expression of a single cr RNA and four cr RNA s. Altogether, our STU CRISPR 2.0 systems further expanded the CRISPR toolbox for plant genome editing and other applications.

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“…With the sgRNA providing targetspecificity, the dCas9-base editors can create point mutations in the target DNA with high specificity and precision. This new technology was quickly adopted in plants, including watermelon (Lu and Zhu 2017;Li et al 2018a;Ren et al 2018;Tian et al 2018). Table 1 summarizes published reports of genome editing in fruit crops.…”

Section: Overview Of Crispr/cas9mentioning

confidence: 99%

Application and future perspective of CRISPR/Cas9 genome editing in fruit crops

Zhou¹,

Wang

et al. 2019

JIPB

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Fruit crops, including apple, orange, grape, banana, strawberry, watermelon, kiwifruit and tomato, not only provide essential nutrients for human life but also contribute to the major agricultural output and economic growth of many countries and regions in the world. Recent advancements in genome editing provides an unprecedented opportunity for the genetic improvement of these agronomically important fruit crops. Here, we summarize recent reports of applying CRISPR/Cas9 to fruit crops, including efforts to reduce disease susceptibility, change plant architecture or flower morphology, improve fruit quality traits, and increase fruit yield. We discuss challenges facing fruit crops as well as new improvements and platforms that could be used to facilitate genome editing in fruit crops, including dCas9‐base‐editing to introduce desirable alleles and heat treatment to increase editing efficiency. In addition, we highlight what we see as potentially revolutionary development ranging from transgene‐free genome editing to de novo domestication of wild relatives. Without doubt, we now see only the beginning of what will eventually be possible with the use of the CRISPR/Cas9 toolkit. Efforts to communicate with the public and an emphasis on the manipulation of consumer‐friendly traits will be critical to facilitate public acceptance of genetically engineered fruits with this new technology.

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Improved Base Editor for Efficiently Inducing Genetic Variations in Rice with CRISPR/Cas9-Guided Hyperactive hAID Mutant

Cited by 181 publications

References 10 publications

Transforming plant biology and breeding with CRISPR/Cas9, Cas12 and Cas13

Transforming plant biology and breeding with CRISPR/Cas9, Cas12 and Cas13

Single transcript unit CRISPR 2.0 systems for robust Cas9 and Cas12a mediated plant genome editing

Application and future perspective of CRISPR/Cas9 genome editing in fruit crops

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