Transformation and Regeneration Protocol for Two Farmer Preferred Open Pollinated Tropical Maize (Zea Mays) Varieties
Journal of Plant Sciences
Volume 4, Issue 4, August 2016, Pages: 80-87
Received: Jul. 5, 2016; Accepted: Jul. 18, 2016; Published: Aug. 3, 2016
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Authors
Johnstone Omukhulu Neondo, Institute for Biotechnology Research, Jomo Kenyatta University of Agriculture and Technology, Nairobi, Kenya
Amos Emitati Alakonya, Institute for Biotechnology Research, Jomo Kenyatta University of Agriculture and Technology, Nairobi, Kenya; Plantain and Banana Improvement Program, International Institute of Tropical Agriculture, Ibadan, Nigeria
Jonathan Matheka, Biotechnology Unit, International Institute of Tropical Agriculture, Nairobi, Kenya; Plant Transformation Laboratory, Department of Biochemistry and Biotechnology, Kenyatta University, Nairobi, Kenya
Joel Okoyo Masanga, Plant Transformation Laboratory, Department of Biochemistry and Biotechnology, Kenyatta University, Nairobi, Kenya
Remmy Wekesa Kasili, Institute for Biotechnology Research, Jomo Kenyatta University of Agriculture and Technology, Nairobi, Kenya
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Abstract
In vitro regeneration of open pollinated varieties (OPVs) Kakamega Striga Tolerant Population 94 (KSTP’94) and ‘Namba Nane’ alongside a tropical inbred line (CML144) was evaluated using immature zygotic embryos as explants. Four callus induction media (CIM) regimes; Murashige and Skoog (MS), Linsmaier and Skoog (LS), Chu (N6) and N6*(N6 medium fortified with 0.35 gL-1 L-proline and 0.8 mgL-1 AgNO3) were evaluated for their potential to induce callus in the three genotypes. All the media were supplemented with sucrose and five levels of 2, 4-Dichlorophenoxyacetic acid (2, 4-D) (0.5, 1.0, 1.5, 2.0 and 2.5 mgL-1). Resulting calli were matured on MS and N6 basal media supplemented with 60 g/L sucrose and similar concentration levels (0.5, 1.0, 1.5, 2.0 and 2.5 mgL-1) of 2, 4-D while the subsequent embryogenic calli were regenerated on hormone-free media. Transformability of these varieties was assessed via histochemical analysis of β-glucuronidase (GUS) reporter gene following Agrobacterium-mediated transformation. Statistical analyses were done using Statistical Analysis Software (SAS) and Graphpad Prism softwares with mean separations achieved at 95% confidence intervals. Of the 2 OPVs, KSTP’94 recorded the highest callus induction frequency (84.4%) while Namba Nane (45.6%) had the lowest. Similarly, KSTP, 94 had the highest mean of mature somatic embryos (59.7%) while Namba Nane recorded the lowest (16.4%). Assessment of regeneration frequencies from embryogenic calli revealed no significant differences among the 3 lines although CML 144 had the highest mean number of juvenile plantlets (36.7%). Analysis of transformation frequency (upon selection of calli on media with basta) showed that Namba Nane recorded the lowest transformation frequency (average 13.5%) some words missing. Transformation frequency (based on GUS positive calli) of these varieties ranged from 0.8 to 2.1%. This work therefore provides an empirical platform for potential introduction of useful genes into these varieties.
Keywords
Callus Induction, Embryogenesis, Plant Regeneration, Open Pollinated Varieties, GUS
To cite this article
Johnstone Omukhulu Neondo, Amos Emitati Alakonya, Jonathan Matheka, Joel Okoyo Masanga, Remmy Wekesa Kasili, Transformation and Regeneration Protocol for Two Farmer Preferred Open Pollinated Tropical Maize (Zea Mays) Varieties, Journal of Plant Sciences. Vol. 4, No. 4, 2016, pp. 80-87. doi: 10.11648/j.jps.20160404.14
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Copyright © 2016 Authors retain the copyright of this article.
This article is an open access article distributed under the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
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