International Journal of Horticulture, 2017, Vol.7, No. 6, 40-46
40
Research Report Open Access
Direct
in
vitro
Regeneration of
Nicotiana plumbaginifolia
L. and the Potential for
Genetic Transformation
Maria Kanwal
1
,
Faiz Ahmad Joyia
1
, Ghulam Mustafa
1
, Muhammad Anjum Zia
2
, Iqrar Ahmad Rana
1
, Muhammad Sarwar Khan
1
1 Centre of Agricultural Biochemistry and Biotechnology (CABB), University of Agriculture Faislabad, Pakistan
2 Depertament of Biochemistry, University of Agriculture Faislabad, Pakistan
Corresponding email
International Journal of Horticulture, 2017, Vol. 7, No. 6 doi
Received: 01 Feb., 2017
Accepted: 02 Mar., 2017
Published: 31 Mar., 2017
Copyright
©2017 Kanwal et al., This is an open access article published under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Preferred citation for this article
:
Kanwal M., Joyia F.A., Mustafa G., Zia M.A., Rana I.A., and Khan M.S., 2017, Direct
in vitro
regeneration of
Nicotiana plumbaginifolia
L. and the potential
for genetic transformation, International Journal of Horticulture, 7(6): 40-46 (doi
Abstract
Wild crop relatives are important source of genetic diversity. Plant tissue culture provides proficienct means of
conservation of endangered wild plants sustaining biodiversity, improving plant traits by virtue of induced somaclonal variation and
trangensis. The objective of study was to establish direct
in vitro
regeneration and biolistic transformation protocol for wild tobacco
N.
plumbaginifolia
using
uidA
gene. For direct
in vitro
regeneration, six combinations of IAA and kinetin were used. Then, biolistic
transformation was carried out using BioRad PDS 1 000 (He) gun and 1 100-psi rupture discs. Transformation was confirmed
through histochemical gus assay. A proficient and reproducible
in vitro
regeneration system was standardized for wild tobacco
named as
Nicotiana
plumbaginifolia
. In present study 1 mg/L kinetin in combination with 0.25 mg/L IAA proved best for direct
in
vitro
shoot regeneration from mature leaf-based explants. Success in direct
in vitro
regeneration circumvented the callogensis step,
accelerating the process of
in vitro
regeneration and increasing the chances of obtaining true to type plants. Individual shoots were
placed on MS medium with/without IBA for rooting. Individual shoots were successfully rooted on MS medium without IBA. Then,
explants were transformed with
uidA
gene using gene gun. Transient assay of bombarded calli showed
gus
activity on histochemical
assay even after a week. Analysis of gus expression has been found as a principal method of plant transformation optimizations.
Keywords
Direct
in vitro
regeneration; Transient gus assay; IAA; Kinetin;
Nicotiana plumbaginifolia
1 Introduction
Wild crop relatives are important source of genetic diversity. They can be helpful for developing new varieties
which are able to withstand challenging environment. Plant breeders have found different ways to use the genetic
diversity of wild relatives to produce new plant varieties (Warschefsky et al., 2014; Brozynska et al., 2015). Genes
from wild relatives have increased the productivity of major crops such as wheat, rice, maize and barley. These
plants have different traits and showed tolerance to salinity, extreme temperature, drought conditions and
resistance to disease (Ford-Lloyd et al., 2011; Porch et al., 2013).
Nicotiana plumbaginifolia
is specie of tobacco
commonly known as Wild Tobacco and Tex-mex Tobacco. It belongs to family (Solanaceae). It is an annual weed
herb with hairy stem. It has height of 60 cm with extending radical and thin leaf branches (Davey et al., 2000).
Tex-Mex Tobacco is native to the American continent. It has great medicinal properties because of antimicrobial
agents. Its dried leaves are used in treatment of several human diseases like rheumatic, swelling in order to relieve
pain (Singh et al., 2010).
Its methanoic extract has been used as folk medicine for treatment of oxidative damage
in liver of chicks (Shah et al., 2016).
By the year 2020, it is estimated that world population will reach to eight billion and food production will have to
be doubled. To meet the challenges of increasing population, it is obligatory to make well use of plant genetic
diversity. Plant genes from wild crops are the primary source of genetic variability for modern crops that are
involved in different traits like higher productivity, exceptional nutritious values and resistance against disease
(Rao, 2004). Genetic diversity refers to the total number of different genetic characteristic in a species. Genetic
diversity is very crucial for species to survive, to adapt towards new environmental conditions and for evolution of
new species. In plants, genetic diversity is important to develop varieties tolerant to biotic as well as abiotic
stresses (Kettenring
et al., 2014).
