Split selectable marker systems utilizing inteins facilitate gene stacking in plants

Guoliang Yuan; Haiwei Lu; Kuntal De; Md Mahmudul Hassan; Yang Liu; Md Torikul Islam; Wellington Muchero; Gerald A. Tuskan; Xiaohan Yang

doi:10.1038/s42003-023-04950-8

Split selectable marker systems utilizing inteins facilitate gene stacking in plants

Guoliang Yuan, Haiwei Lu, Kuntal De, Md Mahmudul Hassan, Yang Liu, Md Torikul Islam, Wellington Muchero, Gerald A. Tuskan, Xiaohan Yang

Research output: Contribution to journal › Article › peer-review

1 Scopus citations

Abstract

The ability to stack multiple genes in plants is of great importance in the development of crops with desirable traits but can be challenging due to limited selectable marker options. Here we establish split selectable marker systems using protein splicing elements called “inteins” for Agrobacterium-mediated co-transformation in plants. First, we show that such a split selectable marker system can be used effectively in plants to reconstitute a visible marker, RUBY, from two non-functional fragments through tobacco leaf infiltration. Next, to determine the general applicability of our split selectable marker systems, we demonstrate the utility of these systems in the model plants Arabidopsis and poplar by successfully stacking two reporters eYGFPuv and RUBY, using split Kanamycin or Hygromycin resistance markers. In conclusion, this method enables robust plant co-transformation, providing a valuable tool for the simultaneous insertion of multiple genes into both herbaceous and woody plants efficiently.

Original language	English
Article number	567
Journal	Communications Biology
Volume	6
Issue number	1
DOIs	https://doi.org/10.1038/s42003-023-04950-8
State	Published - May 2023

Funding

The work was supported by the Center for Bioenergy Innovation, a U.S. Department of Energy (DOE) Bioenergy Research Center supported by the Biological and Environmental Research (BER) program. Oak Ridge National Laboratory is managed by UT-Battelle, LLC for the U.S. Department of Energy under Contract No. DE-AC05-00OR22725. This paper has been authored by UT-Battelle, LLC, under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy. The United States Government retains and the publisher, by accepting the article for publication, acknowledges that the United States Government retains a non-exclusive, paid-up, irrevocable, worldwide license to publish or reproduce the published form of this paper or allow others to do so, for United States Government purposes. The Department of Energy will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan (http://energy.gov/downloads/doe-public-access-plan). The work was supported by the Center for Bioenergy Innovation, a U.S. Department of Energy (DOE) Bioenergy Research Center supported by the Biological and Environmental Research (BER) program. Oak Ridge National Laboratory is managed by UT-Battelle, LLC for the U.S. Department of Energy under Contract No. DE-AC05-00OR22725. This paper has been authored by UT-Battelle, LLC, under Contract No. DE-AC05-00OR22725 with the U.S. Department of Energy. The United States Government retains and the publisher, by accepting the article for publication, acknowledges that the United States Government retains a non-exclusive, paid-up, irrevocable, worldwide license to publish or reproduce the published form of this paper or allow others to do so, for United States Government purposes. The Department of Energy will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan ( http://energy.gov/downloads/doe-public-access-plan ).

Funders	Funder number
DOE Public Access Plan
United States Government
U.S. Department of Energy
Biological and Environmental Research
Oak Ridge National Laboratory	DE-AC05-00OR22725
Center for Bioenergy Innovation

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10.1038/s42003-023-04950-8

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title = "Split selectable marker systems utilizing inteins facilitate gene stacking in plants",

abstract = "The ability to stack multiple genes in plants is of great importance in the development of crops with desirable traits but can be challenging due to limited selectable marker options. Here we establish split selectable marker systems using protein splicing elements called “inteins” for Agrobacterium-mediated co-transformation in plants. First, we show that such a split selectable marker system can be used effectively in plants to reconstitute a visible marker, RUBY, from two non-functional fragments through tobacco leaf infiltration. Next, to determine the general applicability of our split selectable marker systems, we demonstrate the utility of these systems in the model plants Arabidopsis and poplar by successfully stacking two reporters eYGFPuv and RUBY, using split Kanamycin or Hygromycin resistance markers. In conclusion, this method enables robust plant co-transformation, providing a valuable tool for the simultaneous insertion of multiple genes into both herbaceous and woody plants efficiently.",

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AU - Yuan, Guoliang

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AU - De, Kuntal

AU - Hassan, Md Mahmudul

AU - Liu, Yang

AU - Islam, Md Torikul

AU - Muchero, Wellington

AU - Tuskan, Gerald A.

AU - Yang, Xiaohan

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Split selectable marker systems utilizing inteins facilitate gene stacking in plants

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