Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14
Routineproductionoflargenumbersoftransgenicplantsisrequiredtofullyexploitadvancesincassavabiotechnologyandsupportdevelopmentofiprovedgermplasmfordeploymenttofarmers.Thisarticledescribesanimproved,high-efficiencytransformationprotocolforrecalcitrantcassavacultivarTME14preferredinAfrica.Factorsthatfav...
| Main Authors: | , , |
|---|---|
| Format: | Journal Article |
| Language: | Inglés |
| Published: |
Frontiers Media
2015
|
| Subjects: | |
| Online Access: | https://hdl.handle.net/10568/74436 |
| _version_ | 1855541902142603264 |
|---|---|
| author | Nyaboga, E.N. Njiru, J.M. Tripathi, L. |
| author_browse | Njiru, J.M. Nyaboga, E.N. Tripathi, L. |
| author_facet | Nyaboga, E.N. Njiru, J.M. Tripathi, L. |
| author_sort | Nyaboga, E.N. |
| collection | Repository of Agricultural Research Outputs (CGSpace) |
| description | Routineproductionoflargenumbersoftransgenicplantsisrequiredtofullyexploitadvancesincassavabiotechnologyandsupportdevelopmentofiprovedgermplasmfordeploymenttofarmers.Thisarticledescribesanimproved,high-efficiencytransformationprotocolforrecalcitrantcassavacultivarTME14preferredinAfrica.Factorsthatfavorproductionoffriableembryogeniccalli(FEC)werefoundtobeuseofDKWmedium,crushingoforganizedembryogenicstructures(OES)through1–2mmsizedmetalwiremesh,washingofcrushedOEStissuesandshortexposureoftyrosinetosomaticembryos;andtransformationefficiencywasenhancedbyuseoflowAgrobacteriumdensityduringco-cultivation,co-centrifugationofFECwithAgrobacterium,germinationofparamomycinresistantsomaticembryosonmediumcontainingBAPwithgradualincreaseinconcentrationandvariationsofthefrequenyofsubcultureofcotyledonary-stageembryosonshootelongationmedium.Byapplyingtheoptimizedparameters,FECwereproducedforcassavacultivarTME14andtransformedusingAgrobacteriumstrainLBA4404harboringthebinaryvectorpCAMBIA2301.About70–80independenttransgeniclinespermlsettledcellvolume(SCV)ofFECwereregeneratedonselectivemedium.HistochemicalGUSassaysconfirmedtheexpressionofgusAgeneintransformedcalli,somaticembryosandtransgenicplants.ThepresenceandintegrationofthegusAgenewereconfirmedbyPCRandSouthernblotanalysis,respectively.RT-PCRanalysisoftransgenicplantsconfirmedtheexpressionofgusAgene.Thisprotocoldemonstratessignificantlyenhancedtransformationefficiencyoverexistingcassavatransformationprotocolsandcouldbecomeapwerfultoolforfunctionalgenomicsandtransferringnewtraitsintocassava. |
| format | Journal Article |
| id | CGSpace74436 |
| institution | CGIAR Consortium |
| language | Inglés |
| publishDate | 2015 |
| publishDateRange | 2015 |
| publishDateSort | 2015 |
| publisher | Frontiers Media |
| publisherStr | Frontiers Media |
| record_format | dspace |
| spelling | CGSpace744362024-05-01T08:19:57Z Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14 Nyaboga, E.N. Njiru, J.M. Tripathi, L. cassava genetics Routineproductionoflargenumbersoftransgenicplantsisrequiredtofullyexploitadvancesincassavabiotechnologyandsupportdevelopmentofiprovedgermplasmfordeploymenttofarmers.Thisarticledescribesanimproved,high-efficiencytransformationprotocolforrecalcitrantcassavacultivarTME14preferredinAfrica.Factorsthatfavorproductionoffriableembryogeniccalli(FEC)werefoundtobeuseofDKWmedium,crushingoforganizedembryogenicstructures(OES)through1–2mmsizedmetalwiremesh,washingofcrushedOEStissuesandshortexposureoftyrosinetosomaticembryos;andtransformationefficiencywasenhancedbyuseoflowAgrobacteriumdensityduringco-cultivation,co-centrifugationofFECwithAgrobacterium,germinationofparamomycinresistantsomaticembryosonmediumcontainingBAPwithgradualincreaseinconcentrationandvariationsofthefrequenyofsubcultureofcotyledonary-stageembryosonshootelongationmedium.Byapplyingtheoptimizedparameters,FECwereproducedforcassavacultivarTME14andtransformedusingAgrobacteriumstrainLBA4404harboringthebinaryvectorpCAMBIA2301.About70–80independenttransgeniclinespermlsettledcellvolume(SCV)ofFECwereregeneratedonselectivemedium.HistochemicalGUSassaysconfirmedtheexpressionofgusAgeneintransformedcalli,somaticembryosandtransgenicplants.ThepresenceandintegrationofthegusAgenewereconfirmedbyPCRandSouthernblotanalysis,respectively.RT-PCRanalysisoftransgenicplantsconfirmedtheexpressionofgusAgene.Thisprotocoldemonstratessignificantlyenhancedtransformationefficiencyoverexistingcassavatransformationprotocolsandcouldbecomeapwerfultoolforfunctionalgenomicsandtransferringnewtraitsintocassava. 2015 2016-05-25T11:59:27Z 2016-05-25T11:59:27Z Journal Article https://hdl.handle.net/10568/74436 en Open Access application/octet-stream Frontiers Media Nyaboga, E.N., Njiru, J.M., & Tripathi, L. (2015). Factors influencing somatic embryogenesis, regeneration and agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14. Frontiers in Plant Science, 6, 411. |
| spellingShingle | cassava genetics Nyaboga, E.N. Njiru, J.M. Tripathi, L. Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14 |
| title | Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14 |
| title_full | Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14 |
| title_fullStr | Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14 |
| title_full_unstemmed | Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14 |
| title_short | Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14 |
| title_sort | factors influencing somatic embryogenesis regeneration and agrobacterium mediated transformation of cassava manihot esculenta crantz cultivar tme14 |
| topic | cassava genetics |
| url | https://hdl.handle.net/10568/74436 |
| work_keys_str_mv | AT nyabogaen factorsinfluencingsomaticembryogenesisregenerationandagrobacteriummediatedtransformationofcassavamanihotesculentacrantzcultivartme14 AT njirujm factorsinfluencingsomaticembryogenesisregenerationandagrobacteriummediatedtransformationofcassavamanihotesculentacrantzcultivartme14 AT tripathil factorsinfluencingsomaticembryogenesisregenerationandagrobacteriummediatedtransformationofcassavamanihotesculentacrantzcultivartme14 |