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Review
. 2022 Feb 4:4:825042.
doi: 10.3389/fgeed.2022.825042. eCollection 2022.

Mini-Review: Transgenerational CRISPR/Cas9 Gene Editing in Plants

Lennert Impens  1   2 , Thomas B Jacobs  1   2 , Hilde Nelissen  1   2 , Dirk Inzé  1   2 , Laurens Pauwels  1   2
Affiliations
Review

Mini-Review: Transgenerational CRISPR/Cas9 Gene Editing in Plants

Lennert Impens et al. Front Genome Ed. .

Abstract

CRISPR/Cas9 genome editing has been used extensively in a wide variety of plant species. Creation of loss-of-function alleles, promoter variants and mutant collections are a few of the many uses of genome editing. In a typical workflow for sexually reproducing species, plants are generated that contain an integrated CRISPR/Cas9 transgene. After editing of the gene of interest, T-DNA null segregants can be identified in the next generation that contain only the desired edit. However, maintained presence of the CRISPR/Cas9 transgene and continued editing in the subsequent generations offer a range of applications for model plants and crops. In this review, we define transgenerational gene editing (TGE) as the continued editing of CRISPR/Cas9 after a genetic cross. We discuss the concept of TGE, summarize the current main applications, and highlight special cases to illustrate the importance of TGE for plant genome editing research and breeding.

Keywords: CRISPR/Cas9; HI-Edit; egg cell; floral dip; gene editing; pollen.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
Transgenerational gene editing and applications in plants. (A) Principle of transgenerational gene editing (TGE). A transgenic plant represented as a chromosome pair is hemizygous for a CRISPR/Cas9 containing T-DNA locus (red triangle) and edited in both alleles (stars). When crossed with a WT, the resulting progeny either lacks the T-DNA and inherits a single edited allele or inherits the T-DNA, resulting in (transgenerational) editing of the inherited WT allele. (B–D) Examples of TGE. (B) TGE for continued editing of homoeoalleles in wheat. A transgenic line may have edits only in a subset of homoeoalleles at the homologous chromosomes. After self-crossing and selecting plants that inherited the T-DNA, all homoeoalleles may now be edited (Wang et al., 2018b). (C) TGE for allelic variation. In tomato, a loss-of-function mutant (stars) also contains a CRISPR/Cas9 containing T-DNA targeting the promoter of the mutant gene. After a cross with WT, resulting T-DNA containing plants have one loss-of-function allele (star), and an allele with a promoter edit (other symbols). Every individual F1 plant has potentially a different promoter edit and phenotype as the phenotype is not determined by the inherited loss-of-function allele (Rodríguez-Leal et al., 2017). (D) Desired-target mutator (DTM) strategy. A maize plant hemizygous for a CRISPR/Cas9 containing T-DNA locus (red triangle) is crossed with an elite inbred line, resulting in TGE and editing of the elite allele. Additional rounds of TGE and backcrossing result in a new edited variety with no linkage drag (Li et al., 2017a).
FIGURE 2
FIGURE 2
Special cases of TGE. (A) Combining haploid induction and gene editing (HI-Edit) in maize. A WT elite maize inbred line is pollinated using a haploid inducer line that contains a CRISPR/Cas9 containing T-DNA locus (red triangle). After fertilization, the male genome is gradually eliminated, but the temporary presence of CRISPR/Cas9 may still edit the elite allele. After doubling of the haploid plant genome using colchicine, a homozygous edited elite DH0 line is obtained (Kelliher et al., 2019; Wang et al., 2019). (B) CRISPR/Cas9 gene editing in Arabidopsis thaliana using floral dip. The female gametophyte (T0) is transformed using Agrobacterium tumefaciens leading to a transformed egg cell that may already be edited. Self-pollination with WT pollen leads to a fertilized egg cell and subsequent zygote in which TGE may take place.
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