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Examination of Thymic Positive and Negative Selection by Flow Cytometry
Published on: October 8, 2012
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PIG-mediated cassava transformation using positive and negative selection
1Institute for Plant Sciences, ETH-Zentrum / LFW E 17, CH-8092 Zürich, Switzerland e-mail: johanna.puonti-kaerlas@ipw.biol.ethz.ch Fax: +41-1-6321044, , , , , , CH.
Plant Cell Reports
|February 14, 2019
Summary
This study developed new, non-antibiotic selection systems for creating transgenic cassava (Manihot esculenta Crantz) plants. Positive selection using mannose proved effective for regenerating genetically modified cassava.
Area of Science:
- Plant Biotechnology
- Molecular Biology
- Agricultural Science
Background:
- Developing efficient selection systems is crucial for producing transgenic crops like cassava.
- Traditional antibiotic-based selection methods can have limitations.
Purpose of the Study:
- To assess the efficiency of positive selection using mannose and negative selection using hygromycin for transgenic cassava regeneration.
- To optimize selection protocols for producing transgenic cassava plants.
Main Methods:
- Somatic cotyledon and embryogenic suspension explants were used for particle inflow gun bombardment.
- Positive selection with mannose and negative selection with hygromycin were evaluated.
- PCR, RT-PCR, Southern, and northern analyses confirmed stable transformation and transgene expression.
Main Results:
- Transgenic cassava plants were successfully obtained using both mannose-based positive selection and hygromycin-based negative selection (in embryogenic suspensions).
- A novel rooting test on mannose-supplemented medium enhanced the positive selection system's efficacy.
- Stable transformation and transgene expression were rigorously verified.
Conclusions:
- Non-antibiotic positive selection using mannose is a viable method for producing transgenic cassava.
- Optimized selection strategies contribute to more efficient genetic engineering of cassava.
Keywords:
Abbreviations2,4-D 2,4-Dichlorophenoxyacetic acidBA 6-BenzylaminopurineBiolisticsCBM Basic mediumCEM Elongation mediumCIM Embryo induction mediumCMM Maturation mediumCOM Organogenesis mediumCTM Transformation mediumFEC Friable embryogenic callusGD Gresshoff and Doy mediumIBA Indole-3-butyric acidKey words Cassava (Manihot esculenta Crantz)MSM Solid mannose selection mediumMSN FEC Formation and embryo conversion mediumNAAα-Naphthalene acetic acidNegative selectionPositive selectionSCV Settled cell volumeSH Schenk and Hildebrandt mediumSHM Liquid mannose selection mediumTransformation
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