Molecular Plant Breeding 2015, Vol.6, No.16, 1
-
13
9
escape the effects of a control agent that limits the
development and maintenance of the plants (Allendorf
and Luikart, 2007; Maranho et al., 2014), in contrast
to plant populations showing a lower level of
heterozygosity.
The occurrence of null alleles has been pointed out as
a possible problem associated with the use of
microsatellite markers (Callen et al., 1993), which
might result in an individual being scored as homozygote
instead of heterozygous and thus resulting in loss of
information. Our results indicated that out of 70
markers, 19 markers showed less positive values for
the frequency of null alleles, might be regarded as loci
with a possible occurrence of null alleles (O’Reilly
and Wright 1995). On the other hand, the negative
value for the frequency of null alleles (r) observed in
most loci might associate the homozygotes with negative
characters that have been eliminated by humans in
cultivated germplasm (Marinoni et al. 2003). In the
present study most of the markers showed (51 out of
70 ) negative values for the frequency of null allele
occurrence, it indicates that developed markers in the
present study are potential tools for the
differentiation of tobacco species without losing any
information.
In the present study, the probability of identity was
moderately high, which represents the power of each
microsatellite to contribute to a unique genotype for
each species (Dangl et al., 2005). Polymorphic
Information Content (PIC) provides an estimate of
discriminatory power of a marker to differentiate
genotypes based on both the number of alleles
expressed and their relative frequencies (Nagl et al.,
2011). The average of PIC value was 0.57 for perfect
repeats where as for imperfect repeats 0.19, indicates
an isolation of highly polymorphic microsatellites in
Nicotiana
species. These polymorphic markers were
should provide sufficient level of genetic diversity to
investigate the fine scale population structure and
evaluate the breeding strategy (Xu et al., 2008). We
have also used microsatellite markers for revealing
polymorphism among the different
Nicotiana
species
and also assessed for their cross-species transferability.
All the species of
Nicotiana
principally divided based
on the subgenus to which they belong and the number
of chromosomes. The species of subgenus Petunoides
have 9 sections, mostly species belong to the same
section were grouped together. For instance, two
species of Rependae section and two species of Alatae
section were grouped in the same cluster (Ib) with
higher percentage of similarity. Similarly 6 species
of section Suaveolentes used in the present study were
grouped in cluster II. The clustering pattern of the
tobacco species based on the nuclear SSR profile
supports the traditional classification of genus
Nicotiana
with exception of grouping of
N. rustica
and
N. undulate.
Among the subgenus Petunioides,
the section Undulatae and Trigonophyllae are
considered to be close to the subgenera Rustica, so the
species
N. undulata
was grouped along with the
species belongs to the subgenus Rustica as these two
species shares the same number (12) of chromosomes
(Khan and Narayan, 2007) (Figure 3). The microsa-
tellite-based dendrogram was congruent with existing
sectional representations based on morphological,
chromosome and DNA information (Moon et al.,
2008).
In general, the transferability rates observed in this
study were higher than those reported earlier (Moon et
al., 2008), which could be due to high rates of
conservation of primer binding sites. A minimal set
comprising of 11 markers was constituted for the
identification of most of the Italics species. However,
it should be pointed out that only one accession was
used for each species, as the objective was to
demonstrate the feasibility of these markers for
studying genetic variation and relationships at species
level and a detailed study involving more number of
accessions for each species is warranted to validate
these markers. Perfect clustering of genotypes belongs
to the all the eight tobacco types were observed with
these markers. Hence, the type specific markers
identified in the present study for various tobacco
types as well as various species specific markers
belongs to
Nicotiana
would be useful in testing of
purity of different types of tobaccos, studying the
genetic variation among varieties, true hybrid
identification, monitoring introgression of the target
gene(s) and relationships between different tobacco
types within cultivated tobacco using the cultivars
representing a wide range of morphological diversity
present within
N. tabacum
. The devloped markers also
exhibited high PIC values, suggesting their potential
