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Tropical Plant Biology

, Volume 5, Issue 1, pp 127–139 | Cite as

A High-throughput Platform for the Production and Analysis of Transgenic Cassava (Manihot esculenta) Plants

  • Nigel Taylor
  • Eliana Gaitán-Solís
  • Theodore Moll
  • Brent Trauterman
  • Tira Jones
  • Amita Pranjal
  • Cindy Trembley
  • Vince Abernathy
  • David Corbin
  • Claude M. Fauquet
Article

Abstract

A platform for high-throughput production and analysis of transgenic cassava (Manihot esculenta) has been developed for the variety 60444 and implemented to generate plants expressing traits for nutritional enhancement, modified metabolism, promoter analysis and disease resistance. Over a three and a half year period this system has been utilized to produce more than 3500 independent transgenic plant lines from 50 different genetic constructs within a single laboratory. Plants recovered through this system have proven robust and efficacious for engineered traits under greenhouse conditions and within the first confined field trials of transgenic cassava carried out in Uganda, Kenya, Nigeria and Puerto Rico. Detailed procedures are described for the operation of this platform, including all steps in tissue culture, genetic transformation, copy number estimation, greenhouse establishment for shoot and storage root formation and systems for centralized quality control, transgenic plant tracking and regulatory compliance. In addition to providing reliable transgenic plant production for proof of concept studies in the variety 60444, the systems implemented and described here form the structure for high throughput production of transgenic farmer-preferred cultivars of cassava.

Keywords

Cassava DNA genomic dot blot Friable embryogenic callus Genetic transformation 

Abbreviations

BAP

benzylaminopurine

DDPSC

Donald Danforth Plant Science Center

FEC

friable embryogenic callus

GD2

Greshoff and Doy basal medium containing 20 g/l sucrose

GD2 50P

GD2 supplemented with 50 μM picloram

GFP

green fluorescent protein

ILTAB

International Laboratory for Tropical Agricultural Biotechnology

MS2

Murashige and Skoog basal medium containing 20 g/l sucrose

MS2 0.5NAA

MS2 supplemented with 0.5 μM naphthalene acetic acid

MS2 2BAP

MS2 supplemented with 2 μM benzylaminopurine

MS2 50P

MS2 supplemented with 50 μM picloram

MS2 5NAA

MS2 supplemented with 5 μM naphthalene acetic acid

NAA

naphthalene acetic acid

OES

organised embryogenic callus

SCV

settled cell volume

VIRCA

Virus Resistant Cassava for Africa project

Notes

Acknowledgements

The work described was funded by the United States Agency for International Development (USAID) from the people of the United States of America, the Monsanto Fund and The Bill & Melinda Gates Foundation. The authors are appreciative of support provided by Kevin Lutke, DDPSC Plant Tissue Culture and Transformation Facility, Dale Burkhart, Plant Growth Facility and Howard Berg, DDPSC Integrated Microscopy Facility.

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Nigel Taylor
    • 1
  • Eliana Gaitán-Solís
    • 1
  • Theodore Moll
    • 1
  • Brent Trauterman
    • 2
  • Tira Jones
    • 1
  • Amita Pranjal
    • 1
  • Cindy Trembley
    • 1
  • Vince Abernathy
    • 1
  • David Corbin
    • 3
  • Claude M. Fauquet
    • 1
  1. 1.Donald Danforth Plant Science CenterSt. LouisUSA
  2. 2.University of Texas Southwestern Medical CenterDallasUSA
  3. 3.Dow Agro SciencesIndianapolisUSA

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