Tropical maize transformation through cell suspension and protoplast culture systems
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Date
2014-10-03
Authors
Billiah, Moemi
Journal Title
Journal ISSN
Volume Title
Publisher
Kenyatta University
Abstract
Manipulation of tropical maize in tissue culture is crucial to harnessing the benefits of
biotechriology, especially those involving upgrading the genetic base of the maize to resist or
tolerate biotic and abiotic constraints of production. The strategies involved are many, having
transformed from those involving conventional breeding to the more appropriate molecular plant
breeding methods. Transformation technology as a research or breeding tool to improve maize is
routinely used in most industrial and some specialized public laboratories. However,
transformation of many tropical maize inbred lines remains a challenging task, especially when
using Agrobacterium tumefaciens as the transgene delivery method. Availability of very efficient
Agrobacterium-mediated maize inbred line transfonriation protocol will improve future
opportunities for maize genetic manipulation and functional genomics. Most molecular and gene
function studies in maize requiring tissue culture have been done using cell suspensions mainly
among temperate inbred lines of maize in which the protocols have been optimized to suit these
lines. In this study, efficient quick generation of transgenic plants and progeny from five maize
inbred lines will be investigated using an Agrobacterium-mediated standard binary vector system
comparatively with polyethylene glycol (PEG) and calcium phosphate clean gene delivery
systems. These systems areā¢ envisaged to target cell suspension and protoplast cultures as
explants. The cell suspensions and protoplasts developed from the maize inbred lines will be first
pre-screened for regeneration frequency and efficiency on Murashige and Skoog medium. The
established cell suspensions and protoplast culture systems will thus fast-track transformation of
tropical Kenyan adopted white maize using direct. DNA uptake method. The maize
transformation evaluation studies will be done using GUS reporter gene and pmi selectable
marker gene. The transformation efficiency of direct DNA uptake method coupled with the use
of clean transgenes and the new culture systems will be evaluated through histochemical GUS
staining, southern, western blotting and polymerase chain reaction analysis. The established
protocols will be useful in generating transgenic maize lines resistant to various biotic and abiotic stresses.
Description
school of Pure and Applied Sciences