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Development of an intragenic gene transfer and selection protocol for sugarcane resulting in resistance to acetolactate synthase-inhibiting herbicide

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Abstract

Sugarcane is a prime crop for commercial production of bioethanol and table sugar. Genetic engineering is complementing traditional breeding in sugarcane improvement. Costs and delays associated with regulatory approval for a transgenic event are substantial impediments to the commercialization of transgenic crops. Generation of intragenic sugarcane events carrying additional sequences from sexually compatible sorghum resembles traditional plant breeding and may facilitate regulatory approval. Acetolactate synthase (ALS), is a key enzyme in the biosynthetic pathway of branched-chain amino acids, and also a target for ALS-inhibiting herbicides including sulfonylureas and imidazolinones. In this study, we developed an intragenic minimal expression cassette encoding a mutant sorghum acetolactate synthase gene (mALS) under transcriptional control of the sorghum ubiquitin promoter and sorghum HSP 3′UTR. mALS was delivered into tissue cultures of sugarcane cultivar CP 88-1762 by biolistic gene transfer. A suitable selection protocol for recovery of intragenic events was developed for the herbicide bispyribac sodium. Intragenic sugarcane events were identified by PCR, and qRT-PCR analysis revealed events with different levels of mALS expression. Expression of the mALS gene in sugarcane resulted in chlorsulfuron resistance, as demonstrated by lower injury level and increased plant height following application of chlorsulfuron compared to the control cultivar. The results demonstrated that the mALS gene from sorghum supports the production of intragenic, herbicide resistant sugarcane. This is the first report of an intragenic selectable marker for sugarcane.

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Acknowledgments

The authors would like to thank Dr. Hardev Sandhu (Everglades Research and Educational Center, UF-IFAS, Belle Glade, FL) for providing tops of sugarcane cultivar CP 88-1762 and Sun Gro Horticulture, Apopka, FL for donation of the Fafard #2 potting mix.

Authors’ contributions

FA conceived the experiments. FA, RK and JHJ designed the experiments. JHJ constructed the plasmid. HD, RK, YZ, SP and GS generated the intragenic plants, RK, JHJ and HD analyzed the intragenic plants. RK, HD and FA wrote the manuscript. All authors read and approved the final manuscript.

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Authors and Affiliations

  1. Plant Molecular and Cellular Biology Program, Agronomy Department, Genetics Institute, University of Florida, IFAS, Gainesville, FL, USA

    H. Dermawan, R. Karan, J. H. Jung, Y. Zhao, S. Parajuli, G. Sanahuja & F. Altpeter

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  1. H. Dermawan
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  2. R. Karan
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  3. J. H. Jung
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  4. Y. Zhao
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  5. S. Parajuli
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  6. G. Sanahuja
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  7. F. Altpeter
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Correspondence to F. Altpeter.

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Dermawan, H., Karan, R., Jung, J.H. et al. Development of an intragenic gene transfer and selection protocol for sugarcane resulting in resistance to acetolactate synthase-inhibiting herbicide. Plant Cell Tiss Organ Cult 126, 459–468 (2016). https://doi.org/10.1007/s11240-016-1014-5

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  • DOI: https://doi.org/10.1007/s11240-016-1014-5

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