Improved direct transformation via particle bombardment of split-immature embryo explants in soybean (Glycine max)

Chennareddy, Siva; Cicak, Toby; Mall, Tejinder; Effinger, Katherine; Sardesai, Nagesh; Pareddy, D. R.; Sarria, Rodrigo

doi:10.1007/s11240-018-1440-7

Cited by 4 publications

(4 citation statements)

References 52 publications

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“…Direct transformation of immature soybean embryos was performed essentially as described previously (Chennareddy et al ., ). Briefly, pods of soybean cv.…”

Section: Methodsmentioning

confidence: 97%

Zinc finger nuclease‐mediated targeting of multiple transgenes to an endogenous soybean genomic locus via non‐homologous end joining

Bonawitz¹,

Ainley²,

Itaya

et al. 2018

Plant Biotechnology Journal

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Emerging genome editing technologies hold great promise for the improvement of agricultural crops. Several related genome editing methods currently in development utilize engineered, sequence-specific endonucleases to generate DNA double strand breaks (DSBs) at user-specified genomic loci. These DSBs subsequently result in small insertions/deletions (indels), base substitutions or incorporation of exogenous donor sequences at the target site, depending on the application. Targeted mutagenesis in soybean (Glycine max) via non-homologous end joining (NHEJ)-mediated repair of such DSBs has been previously demonstrated with multiple nucleases, as has homology-directed repair (HDR)-mediated integration of a single transgene into target endogenous soybean loci using CRISPR/Cas9. Here we report targeted integration of multiple transgenes into a single soybean locus using a zinc finger nuclease (ZFN). First, we demonstrate targeted integration of biolistically delivered DNA via either HDR or NHEJ to the FATTY ACID DESATURASE 2-1a (FAD2-1a) locus of embryogenic cells in tissue culture. We then describe ZFN- and NHEJ-mediated, targeted integration of two different multigene donors to the FAD2-1a locus of immature embryos. The largest donor delivered was 16.2 kb, carried four transgenes, and was successfully transmitted to T progeny of mature targeted plants obtained via somatic embryogenesis. The insertions in most plants with a targeted, 7.1 kb, NHEJ-integrated donor were perfect or near-perfect, demonstrating that NHEJ is a viable alternative to HDR for gene targeting in soybean. Taken together, these results show that ZFNs can be used to generate fertile transgenic soybean plants with NHEJ-mediated targeted insertions of multigene donors at an endogenous genomic locus.

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“…Direct transformation of immature soybean embryos was performed essentially as described previously (Chennareddy et al ., ). Briefly, pods of soybean cv.…”

Section: Methodsmentioning

confidence: 97%

Zinc finger nuclease‐mediated targeting of multiple transgenes to an endogenous soybean genomic locus via non‐homologous end joining

Bonawitz¹,

Ainley²,

Itaya

et al. 2018

Plant Biotechnology Journal

Self Cite

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“…For example, a TF of up to 6% was achieved with cp4epsps as selectable marker when embryonic axes of mature seeds of the INCASoy-36 Cuban cultivar were bombarded (Soto et al, 2017). Chennareddy et al (2018) combined an immature half-seed explant with an intact embryonic axis, cold and plasmolysis pre-treatment, and a specific somatic embryogenic callus regeneration medium in their protocol. They achieved 5% TF with HPH/hygromycin selection and 2.7% with DSM2/glufosinate selection.…”

Section: Biolistic Transformationmentioning

confidence: 99%

“…Improvement for reducing escapes or chimeric rate has been made when strict select stringency was used, especially some new selectable markers/reagents such as AHAS/imazapyr (Aragão et al, 2000;Rech et al, 2008), EPSPS/glyphosate (Martinell et al, 1999;Guo et al, 2015Guo et al, , 2020aSoto et al, 2017), and AADA/spectinomycin (Martinell et al, 2002). Meanwhile, the modified protocols made use of specific explants, such as somatic embryogenic calli, to reduce the chance of infection with cells at the late development stage, and combined proper selection of chemical agents with high stringency to decrease escape rate dramatically, which led to more than 90% T 0 transgenic plants transmitting their transgenes into T 1 generation (Soto et al, 2017;Chennareddy et al, 2018;Guo et al, 2020a). Therefore,…”

Section: Chimerism In T 0 Transgenic Plantmentioning

confidence: 99%

Progress in Soybean Genetic Transformation Over the Last Decade

Guo

Qiu

et al. 2022

Front. Plant Sci.

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Soybean is one of the important food, feed, and biofuel crops in the world. Soybean genome modification by genetic transformation has been carried out for trait improvement for more than 4 decades. However, compared to other major crops such as rice, soybean is still recalcitrant to genetic transformation, and transgenic soybean production has been hampered by limitations such as low transformation efficiency and genotype specificity, and prolonged and tedious protocols. The primary goal in soybean transformation over the last decade is to achieve high efficiency and genotype flexibility. Soybean transformation has been improved by modifying tissue culture conditions such as selection of explant types, adjustment of culture medium components and choice of selection reagents, as well as better understanding the transformation mechanisms of specific approaches such as Agrobacterium infection. Transgenesis-based breeding of soybean varieties with new traits is now possible by development of improved protocols. In this review, we summarize the developments in soybean genetic transformation to date, especially focusing on the progress made using Agrobacterium-mediated methods and biolistic methods over the past decade. We also discuss current challenges and future directions.

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“…Transformation and regeneration are relatively efficient in rice but the process still takes ~7 months [217,218]. In soybean, it takes 7–9 months when particle bombardment is used for transformation [219]. Efforts to reduce the time required for in vitro cultivation (or to abolish this phase all together) could shorten the overall timelines for the transformation of important crops.…”

Section: Regenerationmentioning

confidence: 99%

Genome Editing in Agriculture: Technical and Practical Considerations

Jansing

Schiermeyer

Schillberg

et al. 2019

IJMS

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The advent of precise genome-editing tools has revolutionized the way we create new plant varieties. Three groups of tools are now available, classified according to their mechanism of action: Programmable sequence-specific nucleases, base-editing enzymes, and oligonucleotides. The corresponding techniques not only lead to different outcomes, but also have implications for the public acceptance and regulatory approval of genome-edited plants. Despite the high efficiency and precision of the tools, there are still major bottlenecks in the generation of new and improved varieties, including the efficient delivery of the genome-editing reagents, the selection of desired events, and the regeneration of intact plants. In this review, we evaluate current delivery and regeneration methods, discuss their suitability for important crop species, and consider the practical aspects of applying the different genome-editing techniques in agriculture.

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Improved direct transformation via particle bombardment of split-immature embryo explants in soybean (Glycine max)

Cited by 4 publications

References 52 publications

Zinc finger nuclease‐mediated targeting of multiple transgenes to an endogenous soybean genomic locus via non‐homologous end joining

Zinc finger nuclease‐mediated targeting of multiple transgenes to an endogenous soybean genomic locus via non‐homologous end joining

Progress in Soybean Genetic Transformation Over the Last Decade

Genome Editing in Agriculture: Technical and Practical Considerations

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