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Efficient production of transgenic cassava using negative and positive selection
P Zhang1, I Potrykus, J Puonti-Kaerlas
1Institute for Plant Sciences, ETH-Zentrum/LFW E 17, Zürich, Switzerland.
Transgenic Research
|February 24, 2001
Summary
Efficiently generate transgenic cassava plants using mannose selection or hygromycin resistance. This study presents novel methods for selecting and regenerating genetically modified cassava, improving transformation efficiency.
Area of Science:
- Plant Biotechnology
- Molecular Biology
- Genetics
Background:
- Cassava (Manihot esculenta Crantz) transformation efficiency needs improvement for genetic enhancement.
- Existing selection systems for plant transformation can be inefficient or rely on microbial antibiotic resistance.
Purpose of the Study:
- To develop and assess efficient selection systems for generating transgenic cassava.
- To compare a positive selection system using mannose with a negative selection system using hygromycin.
Main Methods:
- Regeneration of transgenic cassava from embryogenic suspensions cocultivated with Agrobacterium.
- Utilized mannose (positive selection) and hygromycin resistance (negative selection) for callus line selection.
- Employed GUS assays, PCR, Southern, RT-PCR, and Northern analyses for verification and expression confirmation.
Main Results:
- Achieved high transformation efficiency: 82.6% (mannose) and 100% (hygromycin) of callus lines were transgenic.
- Successfully regenerated 12 morphologically normal transgenic cassava plant lines.
- Developed a mannose-based rooting test as an alternative to GUS assays for eliminating non-transgenic escapes.
Conclusions:
- Transgenic cassava plants can be effectively produced using either antibiotic resistance genes or an antibiotic-free positive selection system.
- The developed methods enhance the efficiency and reliability of cassava genetic transformation.
- Both mannose selection and hygromycin resistance are viable strategies for obtaining transgenic cassava.