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Deng et al., 2011, Ectopic expression of an
AGAMOUS
homolog
NTAG1
from
Chinese narcissus accelerated earlier flowering and senescence in Arabidopsis, Molecular Plant Breeding Vol.2 No.3 (doi: 10.5376/mpb.2011.02.0003)
15
1992). Most of these genes (except
APETALA2
)
belong to MADS-box transcription factors gene
family which control floral organ identity (Melzer et
al., 2009; Saedler et al., 2001; Theissen, 2001). The
C-function gene
AG
plays a central role not only in
specifying sexual organ identity but also in
determining floral meristem termination (Bowman et
al., 1989; Lohmann and Weigel, 2002). In Arabidopsis,
AG
mutation results in the expansion region of the
A
gene class into the center of the flower, which makes
stamens change into petals and carpels into sepals. In
addition,there are additional abnormal flower
produced in the center of
ag
flower. There is
increasing evidence that the role of floral organ
genes is conserved in different plants, although
differences in regulation, redundancy and function of
these genes exist between species (Ferrario et al.,
2006).
Based on the flowering ABCDE model, double flower
formation is commonly used to link to C functional
genes.
NTAG
has been reported being a putative
AG
ortholog cloned from Chinese narcissus only by the
sequence similarity and the expression pattern analysis
(Wang et al., 2006). However, no further functional
analysis was performed to indicate its involvement in
carpel development. In this presrnt paper the isolation
and characterization of
NATG1
gene, an
AG
homolog
from both Chinese narcissus varieties mentioned
above are reported. Sequence and expression pattern
of
NTAG1
gene exhibited the same in both tested
varieties. Furthermore, functional analysis by using
ectopic tests in Arabidopsis showed that
NATG1
might
be involved in the carpel identity and floral transition.
The utilizations of
NATG1
gene in biotechnology are
discussed.
1 Results
1.1 Isolation and characterization analysis of
NTAG1
genes from two narcissus varieties plants
The flowers of Jinzhanyutai consist of five whorls of
organs including three sepals, three petals, a golden
cup-shaped corona, six stamens, and three fused
carpels (Figure 1A). The sepals and petals are white,
extremely similar and known as tepals (Figure 1A).
Yulinglong plants produce metamorphotic flowers in
which stamens are petal-like structure (Figure 1B,
Figure 1B C). In order to study the mechanism of such
double flower development
，
we isolated the
NTAG1
gene, a putative C-function gene with the information
reported by Wang (Wang et al., 2006) from both
narcissus plants respectively
．
Sequence analysis
showed that
NTAG1
genes from both varieties plants
were exactly same. Comparison of the sequence with
that presented by Wang (Wang et al., 2006) showed
that there were 5 bases pair variation between them.
However, there was only one residue changed in the
K-box when the deduced amino acid sequence was
compared (data not shown). The reason of the
sequence difference may be caused by PCR or
resulted from the plant materials got from different
places.
The
NTAG
expression has been shown only in the
third and forth whorl of the flower of Jinzhanyutai by
northern blot (Wang et al., 2006). To further explore
whether its expression changed in Yulinglong,
semi-quantitative RT-PCR was performed and the
results revealed that the expression pattern was nearly
similar in both varieties plants, it was only expressed
in flowers, but not detectable in leaves, stems and
roots (Figure 2A).
1.2 Ectopic expression of
NTAG1
caused early
flowering and affected the floral organ identity in
transgenic Arabidopsis plants
To further investigate whether the sequence and
structure similarity (Wang et al., 2006) is coupled to
the functional similarity between
NTAG1
and C
functional genes, We performed functional analysis
through transgenic plants.
NTAG1
cDNA driven by
cauliflower mosaic virus 35S promoter was therefore
transformed into Arabidopsis plants. Twenty-one
independent transgenic Arabidopsis T
1
plants in
Col
and nine lines in
Ler
ecotype were obtained through
kanamycine screening (results not shown) and PCR
(Figure 2B). Most transformed plants produced nearly
similar and severe transformation in all development
(Figure 1D-J).
NTAG1
overexpressing plants in
Ler
showed reduced size (Figure 1E, compared with 1D),
early flowering, and small and curled leaves with
yellow tips (Figure 1F). Wild type and transgenic
phenotype plants were segregated in T
1
generation. All
transgenic plants flowered significantly early. Some