Abstract
Main conclusion
StLFY-knockout potato plants were developed using CRISPR/Cas9 system. Inflorescences of edited plants transited to flowering, but inflorescence structures lacked flowers and were indeterminate, producing multiple shoot meristems.
Abstract
The tetraploid potato (Solanum tuberosum L.) is an important agricultural crop worldwide. In this study, we used CRISPR/Cas9 to inactivate the potato homolog (StLFY) of the LEAFY gene—a key regulator of the transition to flowering and floral meristem identity—in a tetraploid potato cultivar. We achieved high rates of all-allelic knockouts. Frameshift indels led to phenotypic alterations, including indeterminate inflorescence development and the replacement of flowers with the leafy-like structures.
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Data availability
All relevant data and materials that support the findings of this study are available from the corresponding author upon request.
Abbreviations
- CRISPR/Cas9:
-
Clustered regularly interspaced short palindromic repeat/CRISPR-associated protein 9
- LFY:
-
LEAFY
- NGS:
-
Next-generation sequencing
- sgRNA:
-
Single guide RNA
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Acknowledgements
The work was performed using the scientific equipment of the Center for Collective Use “Biotechnology” at the All-Russia Research Institute of Agricultural Biotechnology (Moscow, Russia; agreement RFMEFI62114×0003). We are grateful to the Laboratory of Electron Microscopy of the Lomonosov Moscow State University Biological Faculty for performing scanning electron microscopy. We strongly appreciate Prof. Khavkin E.E. from All-Russia Research Institute of Agricultural Biotechnology for the research inspiration. We are grateful to the Syntol LLC (Moscow) for NGS-sequencing.
Funding
The work was somehow financially supported by the Comprehensive Research Program “Development of Potato Breeding and Seed Production”.
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425_2022_4032_MOESM1_ESM.pdf
Fig. S1 Potato cultivar Vector first exon sequences of LEAFY gene. Polymorphisms are shaded with gray. sgRNA target sequences are highlighted with green, PAMs are underlined. Target sites of the NGS-primers for editing events detection are highlighted with yellow (PDF 297 KB)
425_2022_4032_MOESM3_ESM.png
Fig. S3 Indel occurrence among potato T0 plants. The number of plants presenting each variant is shown on the y-axis. Bars indicate different sequence variants at the target sites of analyzed plants. Different colors indicate different mutations. wt, wild-type variant. “-” indicates a deletion, “ + ” indicates an insertion. a Lines developed with sgRNA. b Lines developed with sgRNA (PNG 149 KB)
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Fig. S6 Examples of scanning electron microphotographs of the leafy-like structures from different StLFY-knockout potato plants (PNG 15977 KB)
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Fig. S7 Difference in tuber yield between control and T1 LFY-knockout potato plants under lightroom conditions determined with Mann–Whitney U-test. Control sample contains 7 non-transgenic potato cultivar Vector plants. Mutant sample contains 8 T1 plants developed with sgRNA1 and 11 T1 plants developed with sgRNA2. The edited lines developed with different sgRNAs were joined in one sample (Mutant), because there are no any statistical differences in tuber yield between two groups. a LFY-knockout plants have more tubers than control plants. b Total tubers weight of the individual plant is higher for the control plants than for the LFY-knockout plants. c Examples of control tubers. Lines with minimum, medium and maximum tuber number are shown. d Examples of tubers from lines obtained using sgRNA1. Lines with minimum, medium and maximum tuber number are shown. e Examples of tubers from lines obtained using sgRNA2. Lines with minimum, medium and maximum tuber number are shown (PNG 8549 KB)
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Lebedeva, M., Komakhin, R., Konovalova, L. et al. Development of potato (Solanum tuberosum L.) plants with StLEAFY knockout. Planta 256, 116 (2022). https://doi.org/10.1007/s00425-022-04032-9
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DOI: https://doi.org/10.1007/s00425-022-04032-9