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Title: Development and use of a switchgrass (Panicum virgatum L.) transformation pipeline by the BioEnergy Science Center to evaluate plants for reduced cell wall recalcitrance

Abstract

Here, the mission of the BioEnergy Science Center (BESC) was to enable efficient lignocellulosic-based biofuel production. One BESC goal was to decrease poplar and switchgrass biomass recalcitrance to biofuel conversion while not affecting plant growth. A transformation pipeline (TP), to express transgenes or transgene fragments (constructs) in these feedstocks with the goal of understanding and decreasing recalcitrance, was considered essential for this goal. Centralized data storage for access by BESC members and later the public also was essential. A BESC committee was established to codify procedures to evaluate and accept genes into the TP. A laboratory information management system (LIMS) was organized to catalog constructs, plant lines and results from their analyses. One hundred twenty-eight constructs were accepted into the TP for expression in switchgrass in the first 5 years of BESC. Here we provide information on 53 of these constructs and the BESC TP process. Eleven of the constructs could not be cloned into an expression vector for transformation. Of the remaining constructs, 22 modified expression of the gene target. Transgenic lines representing some constructs displayed decreased recalcitrance in the field and publications describing these results are tabulated here. Transcript levels of target genes and detailed wall analyses frommore » transgenic lines expressing six additional tabulated constructs aimed toward modifying expression of genes associated with wall structure (xyloglucan and lignin components) are provided. Altered expression of xyloglucan endotransglucosylase/hydrolases did not modify lignin content in transgenic plants. Simultaneous silencing of two hydroxycinnamoyl CoA:shikimate hydroxycinnamoyl transferases was necessary to decrease G and S lignin monomer and total lignin contents, but this reduced plant growth. A TP to produce plants with decreased recalcitrance and a LIMS for data compilation from these plants were created. While many genes accepted into the TP resulted in transgenic switchgrass without modified lignin or biomass content, a group of genes with potential to improve lignocellulosic biofuel yields was identified. Results from transgenic lines targeting xyloglucan and lignin structure provide examples of the types of information available on switchgrass lines produced within BESC. This report supplies useful information when developing coordinated, large-scale, multi-institutional reverse genetic pipelines to improve crop traits.« less

Authors:
ORCiD logo [1];  [2];  [1];  [3];  [4];  [1];  [2];  [1];  [2];  [2];  [1];  [5];  [4];  [4];  [1];  [1];  [1];  [1];  [1];  [1] more »;  [5];  [4];  [5];  [1];  [2];  [4];  [1];  [1];  [5];  [3] « less
  1. Noble Research Institute, LLC, Ardmore, OK (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  2. Univ. of Tennessee, Knoxville, TN (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  3. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  4. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Univ. of North Texas, Denton, TX (United States)
  5. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Univ. of Georgia, Athens, GA (United States)
Publication Date:
Research Org.:
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1496000
Grant/Contract Number:  
AC05-00OR22725
Resource Type:
Accepted Manuscript
Journal Name:
Biotechnology for Biofuels
Additional Journal Information:
Journal Volume: 10; Journal Issue: 1; Journal ID: ISSN 1754-6834
Publisher:
BioMed Central
Country of Publication:
United States
Language:
English
Subject:
09 BIOMASS FUELS; Reverse genetics; Lignocellulosic feedstocks; Transformation pipeline; Cell wall; Recalcitrance; Lignin; Ethanol; Bioenergy; HCT; XTH

Citation Formats

Nelson, Richard S., Stewart, Jr., C. Neal, Gou, Jiqing, Holladay, Susan, Gallego-Giraldo, Lina, Flanagan, Amy, Mann, David G. J., Hisano, Hiroshi, Wuddineh, Wegi A., Poovaiah, Charleson R., Srivastava, Avinash, Biswal, Ajaya K., Shen, Hui, Escamilla-Treviño, Luis L., Yang, Jiading, Hardin, C. Frank, Nandakumar, Rangaraj, Fu, Chunxiang, Zhang, Jiyi, Xiao, Xirong, Percifield, Ryan, Chen, Fang, Bennetzen, Jeffrey L., Udvardi, Michael, Mazarei, Mitra, Dixon, Richard A., Wang, Zeng -Yu, Tang, Yuhong, Mohnen, Debra, and Davison, Brian H. Development and use of a switchgrass (Panicum virgatum L.) transformation pipeline by the BioEnergy Science Center to evaluate plants for reduced cell wall recalcitrance. United States: N. p., 2017. Web. doi:10.1186/s13068-017-0991-x.
Nelson, Richard S., Stewart, Jr., C. Neal, Gou, Jiqing, Holladay, Susan, Gallego-Giraldo, Lina, Flanagan, Amy, Mann, David G. J., Hisano, Hiroshi, Wuddineh, Wegi A., Poovaiah, Charleson R., Srivastava, Avinash, Biswal, Ajaya K., Shen, Hui, Escamilla-Treviño, Luis L., Yang, Jiading, Hardin, C. Frank, Nandakumar, Rangaraj, Fu, Chunxiang, Zhang, Jiyi, Xiao, Xirong, Percifield, Ryan, Chen, Fang, Bennetzen, Jeffrey L., Udvardi, Michael, Mazarei, Mitra, Dixon, Richard A., Wang, Zeng -Yu, Tang, Yuhong, Mohnen, Debra, & Davison, Brian H. Development and use of a switchgrass (Panicum virgatum L.) transformation pipeline by the BioEnergy Science Center to evaluate plants for reduced cell wall recalcitrance. United States. doi:10.1186/s13068-017-0991-x.
Nelson, Richard S., Stewart, Jr., C. Neal, Gou, Jiqing, Holladay, Susan, Gallego-Giraldo, Lina, Flanagan, Amy, Mann, David G. J., Hisano, Hiroshi, Wuddineh, Wegi A., Poovaiah, Charleson R., Srivastava, Avinash, Biswal, Ajaya K., Shen, Hui, Escamilla-Treviño, Luis L., Yang, Jiading, Hardin, C. Frank, Nandakumar, Rangaraj, Fu, Chunxiang, Zhang, Jiyi, Xiao, Xirong, Percifield, Ryan, Chen, Fang, Bennetzen, Jeffrey L., Udvardi, Michael, Mazarei, Mitra, Dixon, Richard A., Wang, Zeng -Yu, Tang, Yuhong, Mohnen, Debra, and Davison, Brian H. Fri . "Development and use of a switchgrass (Panicum virgatum L.) transformation pipeline by the BioEnergy Science Center to evaluate plants for reduced cell wall recalcitrance". United States. doi:10.1186/s13068-017-0991-x. https://www.osti.gov/servlets/purl/1496000.
@article{osti_1496000,
title = {Development and use of a switchgrass (Panicum virgatum L.) transformation pipeline by the BioEnergy Science Center to evaluate plants for reduced cell wall recalcitrance},
author = {Nelson, Richard S. and Stewart, Jr., C. Neal and Gou, Jiqing and Holladay, Susan and Gallego-Giraldo, Lina and Flanagan, Amy and Mann, David G. J. and Hisano, Hiroshi and Wuddineh, Wegi A. and Poovaiah, Charleson R. and Srivastava, Avinash and Biswal, Ajaya K. and Shen, Hui and Escamilla-Treviño, Luis L. and Yang, Jiading and Hardin, C. Frank and Nandakumar, Rangaraj and Fu, Chunxiang and Zhang, Jiyi and Xiao, Xirong and Percifield, Ryan and Chen, Fang and Bennetzen, Jeffrey L. and Udvardi, Michael and Mazarei, Mitra and Dixon, Richard A. and Wang, Zeng -Yu and Tang, Yuhong and Mohnen, Debra and Davison, Brian H.},
abstractNote = {Here, the mission of the BioEnergy Science Center (BESC) was to enable efficient lignocellulosic-based biofuel production. One BESC goal was to decrease poplar and switchgrass biomass recalcitrance to biofuel conversion while not affecting plant growth. A transformation pipeline (TP), to express transgenes or transgene fragments (constructs) in these feedstocks with the goal of understanding and decreasing recalcitrance, was considered essential for this goal. Centralized data storage for access by BESC members and later the public also was essential. A BESC committee was established to codify procedures to evaluate and accept genes into the TP. A laboratory information management system (LIMS) was organized to catalog constructs, plant lines and results from their analyses. One hundred twenty-eight constructs were accepted into the TP for expression in switchgrass in the first 5 years of BESC. Here we provide information on 53 of these constructs and the BESC TP process. Eleven of the constructs could not be cloned into an expression vector for transformation. Of the remaining constructs, 22 modified expression of the gene target. Transgenic lines representing some constructs displayed decreased recalcitrance in the field and publications describing these results are tabulated here. Transcript levels of target genes and detailed wall analyses from transgenic lines expressing six additional tabulated constructs aimed toward modifying expression of genes associated with wall structure (xyloglucan and lignin components) are provided. Altered expression of xyloglucan endotransglucosylase/hydrolases did not modify lignin content in transgenic plants. Simultaneous silencing of two hydroxycinnamoyl CoA:shikimate hydroxycinnamoyl transferases was necessary to decrease G and S lignin monomer and total lignin contents, but this reduced plant growth. A TP to produce plants with decreased recalcitrance and a LIMS for data compilation from these plants were created. While many genes accepted into the TP resulted in transgenic switchgrass without modified lignin or biomass content, a group of genes with potential to improve lignocellulosic biofuel yields was identified. Results from transgenic lines targeting xyloglucan and lignin structure provide examples of the types of information available on switchgrass lines produced within BESC. This report supplies useful information when developing coordinated, large-scale, multi-institutional reverse genetic pipelines to improve crop traits.},
doi = {10.1186/s13068-017-0991-x},
journal = {Biotechnology for Biofuels},
number = 1,
volume = 10,
place = {United States},
year = {2017},
month = {12}
}

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