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Research article
In vitro propagation of three mosaic disease resistant cassava cultivars
Amitchihoué Franck SESSOU
University of Abomey-Calavi
Corresponding Author
ORCiD: https://orcid.org/0000-0001-9128-6064
License:
This work is licensed under a CC BY 4.0 License. Read Full License
View the published version at BMC Biotechnology.
Background
Cassava is a staple food for over 800 million people globally providing a cheap source of carbohydrate. However, the cultivation of cassava in the country is facing to viral diseases, particularly cassava mosaic disease (CMD) which can cause up to 95% yield losses. With aim to supply farmers demand for clean planting materials, there is need to accelerate the production of the elite cultivars by use of tissue culture in order to cope with the demand.
Methods
Nodal explants harvested from the greenhouse grown plants were sterilised using different concentrations of a commercial bleach JIK (3.85% NaOCl) and varying time intervals. Microshoots induction was evaluated using thidiazuron (TDZ), benzyl amino purine (BAP), and kinetin. Rooting was evaluated using different auxins (Naphthalene acetic acid NAA and Indole-3-butyricacid IBA). PCR-based SSR and SCAR markers were used to verify the presence of CMD2 gene in the regenerated plantlets.
Results
The highest level of sterility in explants (90%) was obtained when 20% Jik was used for 15 minutes. The best cytokinin for microshoots regeneration was found to be kinetin with optimum concentrations of 5, 10 and 20 µM for Agric-rouge, Atinwewe, and Agblehoundo respectively. Medium without growth regulators was the best for rooting the three cultivars. A survival rate of 100%, 98%, and 98% was recorded in the greenhouse for Agric-rouge, Atinwewe, and Agblehoundo respectively and the plantlets appeared to be morphologically normal. The SSR and SCAR analysis of micropropagated plants showed a profile similar to that of the mother plants indicating that the regenerated plantlets retained the CMD2 gene after passing through in vitro culture, as expected with micropropagation.
Conclusion
The nodal explants was established to be 20% of Jik (3.85% NaOCl) with an exposure time of 15 minutes. Kinetin was proved to be the best cytokinins for microshoot formation with the optimum concentration of 5, 10 and 20µM for Agric-rouge, Atinwewe, and Agblehoundo respectively. The protocol developed during this study will be useful for mass propagation of the elite cassava cultivars.
Keywords
cassava mosaic disease, SSR and SCAR markers, In-vitro propagation nodal explant, genetic conformity
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
This is a list of supplementary files associated with this preprint. Click to download.
- S2File.pdf
S2 File. Regenerated plants images underlying the results reported in the Fig 1.
- S4File.pdf
S4 File. Phytosanitary Certificate of Plant material. Includes detailed on the tractability.
- S5File.pdf
S5 File. Mother plants production under greenhouse. Includes detailed on cuttings planting.
- S6File.pdf
S6 File. Acclimatized plantlets underlying the results reported in the Fig 7.
- S1Table.docx
S1 Table. Detailed data on sterilization experiment. Includes the average of clean explants.
- S1Rawdata1.xlsx
Original Data supporting table 1.
- S1RawImages.pdf
S1 Raw - images. Original uncropped images underlying the gel results reported in the Fig 4.
- S2RawData2.xlsx
Original Data analysed to generate table 2.
- S3RawData3.xlsx
Original Data analysed to generate table 3, 4, and 5.
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Received 31 Jul, 2020
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Reviewer #2 agreed
On 26 Jun, 2020
Review #1 received
Received 24 Jun, 2020
Reviewer #1 agreed
On 23 Jun, 2020
Reviewers invited
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This preprint is under consideration at BMC Biotechnology. A preprint is a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Research article
In vitro propagation of three mosaic disease resistant cassava cultivars
Amitchihoué Franck SESSOU
University of Abomey-Calavi
Corresponding Author
ORCiD: https://orcid.org/0000-0001-9128-6064
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Published
Version 4
Posted 17 Sep, 2020
No comments provided
Reviewers invited
Invitations sent on 25 Aug, 2020
Reviewer #1 agreed
On 25 Aug, 2020
Review #1 received
Received 25 Aug, 2020
Editor assigned
On 18 Aug, 2020
Submission checks complete
On 17 Aug, 2020
Editor invited
On 17 Aug, 2020
No comments provided
Editorial decision: Minor revision
On 06 Aug, 2020
Review #2 received
Received 04 Aug, 2020
Reviewers invited
Invitations sent on 31 Jul, 2020
Reviewer #1 agreed
On 31 Jul, 2020
Reviewer #2 agreed
On 31 Jul, 2020
Review #1 received
Received 31 Jul, 2020
Editor assigned
On 12 Jul, 2020
Submission checks complete
On 11 Jul, 2020
Editor invited
On 11 Jul, 2020
No comments provided
Editorial decision: Minor revision
On 02 Jul, 2020
Review #2 received
Received 28 Jun, 2020
Reviewer #2 agreed
On 26 Jun, 2020
Review #1 received
Received 24 Jun, 2020
Reviewer #1 agreed
On 23 Jun, 2020
Reviewers invited
Invitations sent on 22 Jun, 2020
Editor assigned
On 19 Jun, 2020
First submitted
On 18 Jun, 2020
Submission checks complete
On 18 Jun, 2020
Editor invited
On 18 Jun, 2020
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Biotechnology and Bioengineering
License:
This work is licensed under a CC BY 4.0 License. Read Full License
View the published version at BMC Biotechnology.
Background
Cassava is a staple food for over 800 million people globally providing a cheap source of carbohydrate. However, the cultivation of cassava in the country is facing to viral diseases, particularly cassava mosaic disease (CMD) which can cause up to 95% yield losses. With aim to supply farmers demand for clean planting materials, there is need to accelerate the production of the elite cultivars by use of tissue culture in order to cope with the demand.
Methods
Nodal explants harvested from the greenhouse grown plants were sterilised using different concentrations of a commercial bleach JIK (3.85% NaOCl) and varying time intervals. Microshoots induction was evaluated using thidiazuron (TDZ), benzyl amino purine (BAP), and kinetin. Rooting was evaluated using different auxins (Naphthalene acetic acid NAA and Indole-3-butyricacid IBA). PCR-based SSR and SCAR markers were used to verify the presence of CMD2 gene in the regenerated plantlets.
Results
The highest level of sterility in explants (90%) was obtained when 20% Jik was used for 15 minutes. The best cytokinin for microshoots regeneration was found to be kinetin with optimum concentrations of 5, 10 and 20 µM for Agric-rouge, Atinwewe, and Agblehoundo respectively. Medium without growth regulators was the best for rooting the three cultivars. A survival rate of 100%, 98%, and 98% was recorded in the greenhouse for Agric-rouge, Atinwewe, and Agblehoundo respectively and the plantlets appeared to be morphologically normal. The SSR and SCAR analysis of micropropagated plants showed a profile similar to that of the mother plants indicating that the regenerated plantlets retained the CMD2 gene after passing through in vitro culture, as expected with micropropagation.
Conclusion
The nodal explants was established to be 20% of Jik (3.85% NaOCl) with an exposure time of 15 minutes. Kinetin was proved to be the best cytokinins for microshoot formation with the optimum concentration of 5, 10 and 20µM for Agric-rouge, Atinwewe, and Agblehoundo respectively. The protocol developed during this study will be useful for mass propagation of the elite cassava cultivars.
Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7