These approaches, collectively known as T-DNA-free genome editing, allow scientists to modify crop traits while avoiding the persistent transgenic sequences often associated with conventional CRISPR methods. By summarizing multiple innovative strategies—from transient gene-editing systems to virus- and pollen-mediated editing—the researchers highlight how these technologies can accelerate crop breeding while reducing regulatory and biosafety concerns.
Genome editing has transformed plant biology by enabling precise DNA modifications for functional genomics and crop improvement. However, conventional genome-editing systems—especially CRISPR/Cas delivered through Agrobacterium-mediated transformation or particle bombardment—often introduce foreign DNA elements such as Cas9 or selectable markers into the plant genome. These inserted sequences, known as T-DNA, may persist after editing, raising biosafety concerns and increasing the risk of unintended genetic effects. Eliminating these transgenes typically requires several generations of breeding, a process that is slow and sometimes impractical for crops with long life cycles or those propagated asexually, including potatoes and strawberries. These challenges have driven the development of genome-editing strategies that generate edited plants without integrating foreign DNA.
A review (DOI:10.48130/abd-0026-0001) published in Agrobiodiversity on 25 February 2026 by Yang Li’s team, Biorun Biosciences Co. Ltd, underscores the potential of T-DNA-free editing to enable faster development of improved crops with enhanced productivity, resilience, and sustainability.
This review highlights multiple strategies—including visual screening systems, Transgene Killer CRISPR (TKC), transient DNA or RNA expression, ribonucleoprotein delivery, virus-mediated editing, pollen-mediated editing, and graft-mobile systems—that enable precise genome modification in plants while avoiding stable integration of foreign DNA. Visual screening approaches rely on easily detectable markers, such as fluorescent proteins or pigment-producing genes, allowing researchers to rapidly distinguish edited plants from those carrying transgenic sequences and thereby reducing the need for extensive molecular testing. Another key strategy is the Transgene Killer CRISPR system, which integrates genetic “suicide cassettes” designed to eliminate transgenes during plant reproduction, ensuring that only edited, transgene-free offspring remain; improved versions incorporate visual reporters to further reduce the chance of transgene escape. Complementing these genetic self-elimination systems, transient expression technologies introduce CRISPR/Cas components into plant cells in the form of DNA or RNA that functions only temporarily before being degraded. Because these editing components do not integrate into the genome, regenerated plants can carry desired mutations without retaining foreign DNA. Similarly, ribonucleoprotein-mediated editing delivers preassembled Cas proteins and guide RNAs directly into plant cells, enabling rapid genome editing while minimizing off-target effects. Additional delivery strategies expand the applicability of genome editing to species that are difficult to transform through conventional tissue culture. Virus-mediated editing employs plant viruses to transport CRISPR components throughout plant tissues, enabling gene modification without stable transformation. Pollen-mediated editing uses haploid-inducer pollen to introduce editing machinery during fertilization, facilitating the rapid production of homozygous edited plants. Meanwhile, graft-mobile editing allows gene-editing signals to move from transgenic rootstocks to non-transgenic shoots, generating heritable genome modifications without introducing foreign DNA into the final plant. Together, these strategies significantly expand the genome-editing toolbox and broaden the range of crops that can benefit from advanced molecular breeding technologies.
In summary, this review emphasizes that T-DNA-free genome editing represents a major step toward safer and more efficient crop biotechnology. By avoiding stable transgene integration, these technologies reduce biosafety concerns, simplify regulatory pathways, and increase public acceptance of gene-edited crops. Although challenges remain—including delivery efficiency, regeneration difficulties, and ensuring heritable edits—continued innovation in editing systems and delivery strategies is expected to make T-DNA-free genome editing a central tool for future crop improvement and global food security.
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References
DOI
10.48130/abd-0026-0001
Original Source URL
https://doi.org/10.48130/abd-0026-0001
About Agrobiodiversity
Agrobiodiversity is the official journal of Yunnan Agricultural University and published by Maximum Academic Press. Agrobiodiversity is an open access, online-only, rigorously peer-reviewed academic journal focusing on the research and studies related to agriculture and biodiversity, including but not limited to: innovation discovery, theory, and technology of agricultural biodiversity; diversity of agricultural genetic resources; environmental interactions among various crops; interaction between crops and abiotic environment; interaction between crops and microbial environment; research on new composite agricultural technology; exploration of new resource species in agriculture. Agrobiodiversity is dedicated to publishing original research articles, reviews, perspectives, opinions, letters, and editorials with high quality.