International Research Journal of Plant Science (ISSN: 2141-5447) Vol. 4(5) pp. 109-116, May, 2013
Available online http://www.interesjournals.org/IRJPS
Copyright © 2013 International Research Journals
Full Length Research Paper
Molecular Cloning of Senescence-Related cDNA,
OsRab7, from Thai Jasmine Rice
(Oryza sativa L. cv. KDML 105)
Sugunya Pitakrattananukool1, Supranee Sitthiphrom2, Robert W. Cutler3,
Somboon Anuntalabhochai1,4*
*1
Department of Biology, Faculty of Science, Chiang Mai University, Muang, Chiang Mai 50200, Thailand.
2
Faculty of Science and Technology, Loei Rajabhat University, Loei, 42000, Thailand.
3
Program of Physics, Department of Science, Edison State College, Ft. Myers, Florida, 33919, USA.
4
* Biotechnology Unit, University of Phayao, Muang, Phayao, 56000, Thailand.
*Corresponding Author E-mail: soanu.1@gmail.com
Abstract
In this work we cloned an OsRab7 DNA sequence encoding a small GTP-binding protein from Thai
jasmine rice (Oryza sativa L. cv. KDML 105). All of the characteristic motifs of the small GTP-binding
Rab family proteins were present in the OsRab7 sequence. The expression analysis revealed that this
cDNA was present in various rice tissues including roots, young leaves, senescing leaves, flowers and
seeds. In addition, the cDNA was also up-regulated under the following induced-senescence
conditions, abscisic acid (ABA), ethylene, NaCl, dark treatment and wounding, suggesting that OsRab7
acted as a stress-inducible gene involved in the senescence process in Thai jasmine rice.
Key words: Small GTP-binding protein, OsRab7, stress-inducible gene, leaf senescence
INTRODUCTION
The small GTP-binding proteins are known as monomeric
G proteins with molecular masses of 20-40 kDa. Several
lines of evidence indicate that the small GTP-binding
protein existed in eukaryotes and regulated diverse
processes in eukaryotic cells including cell proliferation,
signal transduction, vesicular transport, cytoskeleton
organization and intracellular membrane trafficking. Their
molecular function was to switch cycling between the
guanosine triphosphate (GTP) activated and guanosine
diphosphate (GDP) inactivated state (Bischoff et al.,
1999; Ma, 2007). In particular the Rab family contains a
large number of proteins which play an important role in
various intracellular trafficking pathways in plants
(Agarwal et al., 2009). This family is divided into 8
subfamilies designated A-H for each individual class
(Vernoud et al., 2003). Most Rab GTPase are
approximately 24 kDa in molecular weight containing
about 220 amino acids. A total of five Rab subfamily
regions (RabSF) and four Rab family regions (RabF)
have been described, where these designations
correspond to the surface loops involved in proteinprotein interactions. The hypervariable region at the C-
terminus called the dicysteinyl prenylation signal is
indicated by a ‘CC’ (Pereira-Leal and Seabra, 2001).
Several cDNA regions encoding Rab proteins have
been cloned from various plant species and their
functions have been characterized. For example, the
Rha1(Rab5) region was shown to play a role in vascular
trafficking in the root tip and stomal guard cells
(Anuntalabhochai et al., 1991; Terryn et al., 1993; Sohn
et al., 2003). Rab5a was shown to be involved in
vesicular membrane transport and also participate in the
transport of proglutelin from the Golgi to the PSV.
Furthermore, Rab5a is required for the maintenance of
the general structural organization of the endomembrane
system in developing rice seeds (Fukuda et al., 2011).
The transcript expression of the PgRab7 gene isolated
from Pennisetum glaucum was found to be differentially
up-regulated by such environmental stimuli as cold,
dehydration and NaCl and also by the plant hormone
IAA. Overexpression of the PgRab7 gene enhanced
tolerance to NaCl and mannitol in transgenic tobacco
(Agarwal et al., 2008). Moreover, Vanlandingham
and Ceresa (2009) reported that RAB7 is required for the
110 Int. Res. J. Plant Sci.
transfer of cargo from the late endosome/multivesicular
body to the lysosome and for endocytic organelle
maintenance.
Leaf senescence is the final stage of leaf
development and involves programed cell death. The rice
senescence process is similar to that of other plants and
starts from the lower leaves before extending upward as
the plant grows with spreading leaf yellowing. This
significant change in cell structure comes from the
breakdown of the chloroplast and degradation of various
macromolecules. The degraded products are then
transferred to sink organs as recaptured nutrients (Lim et
al., 2007). From molecular studies during senescence,
many genes either have gradually increasing expression
levels or are completely switched off. Moreover, many
new transcripts were shown to have new functions during
senescence. Recently, some members of the Rab genes
were reported as being involved in leaf senescence. For
example, Price et al. (2008) determined that some Rab
subfamily genes were upregulated in senescent of the
wallflower petal, and RabG3b was found to be a
modulator for cell death progression during the pathogen
response and senescence process in Arabidopsis (Kwon
et al., 2009).
In this work we demonstrate that the OsRab7 gene is
related to senescence in Thai jasmine rice and
characterize the function and sequence of this member of
the small GTP-binding proteins.
MATERIALS AND METHODS
Plant Preparation
Thai jasmine rice (Oryza sativa L. cv. KDML 105) seeds
were germinated on agar containing MS media
(Murashige and Skoog, 1962) under continuous light at
o
25 C. After 10 days of germination, the roots were
collected from some rice seedlings and other rice
seedlings were then transferred into soil in a greenhouse.
The third leaf blades were collected at 20, 30, 40 and 50
days after germination (DAG), in addition to the flowers
and seeds at 15 days after flowering (DAF).
Plant treatment and Induced-senescence Condition
For the plant growth regulator treatments, the third leaf
samples were chopped to about 2 to 3 cm in length
segments. These chopped rice leaves were then
incubated in 3 mM MES buffer at a pH of 5.8
supplemented with 1 mM ethephon and 100 µM
of abscisic acid (ABA) under continuous light to
examine the effect of these growth regulators. The
leaf samples were taken at 0, 3, 6, 12 and 24 h
respectively.
For the salinity treatments, the chopped rice leaves were
incubated in 3 mM MES buffer pH 5.8 supplemented
with 100 mM and 200 mM NaCl under continuous light.
The leaf samples were taken at 0, 3, and 5 days
respectively.
For the dark treatment, the chopped leaves were
incubated in 3mM MES buffer pH 5.8 and kept in
darkness and incubated for 0, 3 and 5 days.
For the wounding treatment, the third leaf samples of
the rice seedlings at 15 DAG were wounded using a
needle stab. Leaf samples were then taken at 0, 3 and 5
days after wounding.
Measurement of Chlorophyll Content
2
Chlorophyll contents were measured for 0.6 x 0.6 cm
rice leaf samples. The tissues were weighed and ground
in liquid nitrogen, and then extracted with 100%
methanol. The absorbance was measured at 652.0 nm
and 665.2 nm. The chlorophyll concentration per fresh
weight was then calculated using the equation described
by Porra (2002).
chlorophyll (a+b) = 22.12(A652.0) + 2.71(A665.2)
Cloning of a Full-length OsRab7 cDNA from Thai
Jasmine Rice
The partial cDNA sequences encoding the OsRab7
protein were obtained using PCR amplification and total
RNA was extracted from the rice leaves at 40 DAG using
Trizol reagent (Invitrogen). First-strand cDNA was then
synthesized using Superscript III Reverse transcriptase
(Invitrogen) following the manufacturer’s instructions. The
first-strand cDNA was used as the template for the PCR
amplification. The degenerate forward primer was
designed to correspond to the highly conserved Nterminal region of the small GTP-binding proteins. The
forward
primer
(OsRabFD),
5’GGNGAYNHNGSNRYNGGNAAR-3’, was constructed
to match a sequence coding for the highly conserved
2+
phosphate/Mg
binding domain; DTAGQE(RKS). A
poly-A oligonucleotide was also used as a reverse
primer, oligo(dA)18; 5’-AAAAAAAAAAAAAAAA AA-3’. The
amplification conditions were as follows, an initial
o
denaturation at 94 C for 2 min, followed by 35 cycles at
o
o
94 C for 45 s, annealing at 50 C for 30 s, extension at 72
o
o
C for 1 min, and a final extension at 72 C for 10 min.
The DNA fragments were ligated into the pTZ57R vector
(Fermentas). DNA sequencing was performed using the
dideoxynucleotide chain termination method (Sanger et
al., 1997) using an automated sequencer (ABI). The fulllength of the OsRab7 cDNA sequences were obtained
using by 5’ Full Core Set (Takara) following the
manufacturer’s instructions.
Pitakrattananukool et al. 111
Semi Quantitative
(QPCR) Reaction
Reverse
Transcriptase
PCR
Total RNA was extracted from the rice tissues using
Trizol reagent and first-strand cDNA was synthesized
using a Superscript III Reverse transcriptase (Invitrogen)
according to the manufacturer’s instructions. Fifteen ng of
first-strand cDNA was used as the PCR template. The
amplified PCR product sizes were in the 200 - 500 bp
range. The QPCR reaction was performed as follows:
o
initiation for template denaturation for 2 min at 94 C
o
followed by 26 cycles of denaturation for 30 s at 94 C,
o
o
annealing for 30 s at 62 C and extension for 30 s at 72 C.
Rice β-Actin transcripts were used as the internal
standards.
QPCR Analysis
Agarose electrophoresis was performed to visualize the
PCR products and photographs were taken. The
intensities of the bands were analyzed using Scion Image
software (Scion Corporation, Frederick, Maryland) to
monitor the relative level of gene expression. The level of
transcript expression was presented as the ratio of
expression of OsRab7/β-Actin.
RESULTS AND DISCUSSION
Cloning Of Full-Length cDNAs Encoding Small GTPBinding Rab Family Proteins From Thai Jasmine Rice
We attempted to isolate cDNA transcripts encoding small
members of the GTP-binding Rab family from Thai
jasmine rice. First, the partial cDNAs were amplified
using QPCR. The amplification was performed using a
degenerate forward primer corresponding to the highly
2+
conserved phosphate/Mg binding domain and a specific
reverse primer; oligo(dA)18. A DNA fragment
approximately 770 bp in length was obtained and cloned
into a pTZ57R vector. This fragment was sequenced and
the amino acid sequences were predicted from the
nucleotide sequence. The protein sequence representing
the -CC- motif (the geranylgeranylation region) at the Cterminal was found to be present in this Rab sequence.
After the entire sequence was cloned and analyzed, this
cDNA fragment was found to belong to the Rab7 family for
japonica rice and was designed as OsRab7.
The nucleotide and deduced protein sequences of
OsRab7 are shown in Figure 1. This cDNA is 1,010 bp in
length and contains 65 bp of 5’-untranslated region, an
open reading frame of 628 bp, including 317 bp of a 3’untranslated region (excluding the poly-A tail). The ATG
codon was located at position 66 and the TGA codon at
position 691. The polypeptide of 206 amino acids had a
calculated molecular mass of 23 kDa.
Characterization
Sequences
of
the
Deduced
Amino
Acid
A homology search was done using Blast which found
that the deduced amino acid sequence of OsRab7 had
strong homology to many small GTP-binding proteins
(Rab7) in the Rab family from various plant species. The
sequence was also aligned with ClustalW (Altschul et al.,
1990) which showed that the sequence of Rab7 showed
high homology to Rab7 from other plants [97% to Oryza
sativa (japonica cultivar group), 90% to Cucumis sativus,
89% to Arabidopsis thaliana, and 88% to Lotus
japonicus], with the highest score being to Rab7 from
Oryza sativa (indica cultivar group) (98% identity). The
multiple alignment result also revealed that the conserved
region, including the Rab specific motif (RF), Rab
subfamily
regions
(RSF),
the
guanine
and
2+
phosphate/Mg binding site (G and PM) and double
cysteine C-terminal motif were all contained in the
OsRab7 sequence (Figure 2).
Expression Profile Analysis of Osrab7 in Thai
Jasmine Rice Tissues
The expression of OsRab7 was examined using QPCR.
The results revealed that OsRab7 was expressed in all
rice tissues: roots, leaves, flowers and seeds [Figure.
3(A)]. The relative level of expression was analyzed from
the intensity of bands using the Scion Image program
[Figure. 3(B)].
From Figure. 3. the expression of OsRab7 was
monitored in all rice tissues tested indicating that OsRab7
plays a role in several development processes in Thai
jasmine rice. However, the results revealed an
approximately 3 fold up-regulation of OsRab7 mRNA in
50 DAG leaves as opposed to senescing leaves than in
20 DAG leaves or young leaves. Thus, the expression of
OsRab7 was examined in leaves treated with stressinducible senescence conditions to find out whether
OsRab7 was involved in the senescence process in Thai
jasmine rice.
Expression Profile Analysis of Osrab7 Under Stress
Induced-Senescence Conditions
The Expression of OsRab7 was determined in Thai
jasmine rice leaves under induced-stress conditions. The
QPCR reactions revealed that the expression of OsRab7
was 2 - 6 fold up-regulated in the rice leaves exposed to
100 and 200 mM of NaCl for 3 and 5 days, 100 µM of
ABA which showed the highest expression at 12 h, and
kept in darkness for 3 and 5 days and for wounded
leaves. Whereas, it was only slightly detected in leaves
exposed to 1 mM of ethephon (Figure 4).
112 Int. Res. J. Plant Sci.
1
acctcGtcttcccgttccccgcgccgcgcgggctcgctccccgcgggggcagcttctaga
61
tcccgATGGCCTCCCGCCGCCGCACCCTACTCAAGGTCATCATCCTGGGCGACCCGGGGG
M
121
A
S
R
R
R
T
L
L
K
V
I
I
L
G
D
P
G
V
TTGGGAAGACGTCCCTGATGAACCAATATGTGAACAAGAAGTTCAGCAACCAGTACAAGG
G
K
T
S
L
M
N
Q
Y
V
N
K
K
F
S
N
Q
Y
K
A
181
CTACGATTGGCGCGGATTTCCTCACCAAGGAGGTTCAGTTCGAGGATAGGCTCTTCACTT
241
TGCAAATATGGGATACTGCTGGCCAGGAAAGGTTTCAGAGTCTTGGTGTTGCATTCTACC
T
Q
301
I
G
W
A
D
D
T
F
A
L
G
T
Q
K
E
E
R
V
F
Q
Q
F
S
E
L
D
G
R
V
L
A
F
F
T
Y
L
R
79
A
D
C
C
V
L
V
Y
D
V
N
S
M
K
S
F
D
N
L
99
N
W
R
E
E
F
L
I
Q
A
S
P
S
D
P
D
N
F
P
119
CTTTTGTTCTTTTGGGCAACAAAGTTGATGTAGACAGTGGCAACAGCCGTGTGGTCTCTG
F
V
L
L
G
N
K
V
D
V
D
S
G
N
S
R
V
V
S
E
481
AGAAGAAGGCAAAGGCATGGTGTGCCTCTAAAGGGAATATCCCATACTTTGAGACATCTG
541
CCAAGGATGGTACAAACGTGGAGGAGGCTTTCCAGTGCATTGTAAAGAATGCTCTGAAGA
K
K
601
59
TTAACAACTGGCGTGAAGAATTTCTAATTCAGGCAAGCCCATCAGACCCTGATAACTTCC
N
421
I
39
GTGGAGCAGATTGCTGTGTTCTAGTTTATGATGTCAATTCTATGAAGTCATTTGATAATC
G
361
19
K
D
A
G
K
T
A
N
W
V
C
E
A
E
S
A
K
F
G
Q
N
C
I
I
P
V
Y
K
F
N
E
A
T
L
S
K
A
N
139
159
179
ATGAACCAGAGGAAGAACTGTATGTGCCGGACACCGTGGATGTGGTGGGTGGCAACCGGC
E
P
E
E
E
L
Y
V
P
D
T
V
D
V
V
G
G
N
R
P
661
CCCCAAGATCATCCCGCTGCTGCTAGgacgtgatggaccatgaggggccagactgttggc
721
tatgcggtaacagaactacctttccacattgctgtgccaccatggtacctctcaaggacc
781
cattcgtaaccttttcaatcacctcatgtacccaattaagattgatgcgtctggcctgag
841
ttgtcaaatttgtggatgttgtgcaatttaggggtagcgtcatatctttgtgaatacaat
901
cggtgaaataagatgagtgtaaactgaagtttctccattatggttctctctgaaacgaac
961
aagatgaaattgttctgtctgcattgaggctgaaaaaaaaaaaaaaaaaa
P
R
S
S
R
C
C
Stop
199
206
Figure 1. The nucleotide sequence and deduced amino acid sequence of OsRab7. Bold letters
indicate the start and stop codons, while the 5’ and 3’ UTR are indicated by lower case letters. The
predicted amino acid sequence is shown below the nucleotide sequence in single-letter code. The CC- motif is underlined. The primer binding site and direction to amplify the partial cDNA is indicated
by the arrows.
Stress Induced-Senescence Conditions Reduced
Chlorophyll Content in Thai Jasmine Rice Leaves
The chlorophyll content in the leaf tissues was one
indicator of the senescence parameter. Stress conditions
such as NaCl, ABA, ethephon, darkess and wounding
were chosen to induce a decrease in chlorophyll content
of the rice leaves. The chopped leaf samples under
stress conditions were then used to determine the
chlorophyll content which is a parameter related to the
senescence process. The chlorophyll content remaining
in the leaf samples under stress conditions were then
measured using the method and analysis first described
by Porra (2002). The results showed that the chlorophyll
content decreased in leaves (Figure 5) with the highest
expression levels of OsRab7 as is shown in Figure 3.
Expression Profile Analysis of Osrab7 in Thai
Jasmine Rice Leaves Under Natural Senescence
Conditions
Since OsRab7 showed higher expression in senescing
leaves than in young leaves (Figure 3.) and was
Pitakrattananukool et al. 113
LoRab7
OsRab7
Os_inRab7
Os_jaRab7
CuRab7b
AtRab71
LoRab7
OsRab7
Os_inRab7
Os_jaRab7
CuRab7b
AtRab71
LoRab7
OsRab7
Os_inRab7
Os_jaRab7
CuRab7b
AtRab71
LoRab7d
OsRab7
Os_inRab7
Os_jaRab7
CuRab7b
AtRab71
G1 RF1
PM1
RSF2 PM2
RF2
RSF1
MASRRRMLLKVIILGDSGVGKTSLMNQYVNRKFSNQYKATIGADFLTKEVQFEDRLFTLQ
MASRRRTLLKVIILGDPGVGKTSLMNQYVNKKFSNQYKATIGADFLTKEVQFEDRLFTLQ
MASRRRTLLKVIILGDTGVGKTSLMNQYVNKKFSNQYKATIGADFLTKEVQFEDRLFTLQ
MASRRRTLLKVIILGDSGVGKTSLMNQYVNKKFSNQYKATIGADFLTKEVQFEDRLFTLQ
MPSRRRTLLKVIILGDSGVGKTSLMNQYVNKKFSNQYKATIGADFLTKEVQFEDRLFTLQ
MPSRRRTLLKVIILGDSGVGKTSLMNQYVNKKFSNQYKATIGADFLTKEVQFEDRLFTLQ
*.:*** *********.*************:********************::*******
PM3
RF3
RF4
IWDTAGQERFQSLGVAFYRGADCCVLVYDVNVMKSFDNLNHWREEFLIQASPSDPENFPF
IWDTAGQERFQSLGVAFYRGADCCVLVYDVNSMKSFDNLNNWREEFLIQASPSDPDNFPF
IWDTAGQERFQSLGVAFYRGADCCVLVYDVNSMKSFDNLNNWREEFLIQASPSDPDNFPF
IWDTAGQERFQSLGVAFYRGADCCVLVYDVNSMKSFDNLNNWREEFLIQASPSDPDNFPF
IWDTAGQERFQSLGVAFYRGADCCVLVYDVNSMKSFDNLNNWREEFLIQASPSDPENFPF
IWDTAGQERFQSLGVAFYRGADCCVLVYDVNSMKSFENLNNWREEFLIQASPSDPENFPF
******************************* ***::**:******:*******:****
VVLGNKIDVDGGNSRVISEKKAKAWCASKGNIPYFETSAKEGFNVEAAFQCIAKNALKNE
VLLGNKVDVDSGNSRVVSEKKAKAWCASKGNIPYFETSAKDGTNVEEAFQCIVKNALKNE
VLLGNKVDVDSGNSRVVSEKKAKAWCASKGNIPYFETSAKDGTNVEEAFQCIVKNALKNE
VLLGNKVDVDGGNSRVVSEKKAKAWCASKGNIPYFETSAKDGTNVEEAFQCIVKNALKNE
VVLGNKVDVDGGNSRVVSEKKARAWCASKGNIPYFETSAKEGINVEEAFQCIAKNALKSG
VLIGNKVDVDDGNSRVVSEKKAKAWCASKGNIPYFETSAKVGTNVEEAFQCIAKDALKSG
*::***:***.****.:*****:***************** * *** **:**.::*:*.
C
PEEEMYLPDTIDVGGGGRQQRSTGCEC 207
PEEELYVPDTVDVVGGNRPPRSSRCC- 206
PEEELYVPDTVDVVGGNRAPRSSGCC- 206
PEEELYVPDTVDVVGGNRAQRSSGCC- 206
EEEEIYLPDTIDVGSNNQ-PRSSGCDC 206
EEEELYLPDTIDVGTSNQ-QRSTGCEC 206
*:::*:***:**
.: * : *
60
60
60
60
60
60
120
120
120
120
120
120
180
180
180
180
180
180
Figure 2. Alignment of the deduced OsRab7 amino acid sequence from jasmine rice. Distinct functional
domains are designed according to Pereira-Leal and Seabra (2001). Rab specific regions (RF), Rab
subfamily specific regions (RSF), GDP/GTP-binding domains (G), Phosphate/M2+ binding domains (PM),
and the geranylgeranylation region (C) are surrounded by boxes. The amino acid sequences were obtained
from Oryza sativa [indica cultivar group] (Os_in), Oryza sativa [japonica cultivar group] (Os_ja), Cucumis
sativus (Cu), Arabidopsis thaliana (At), and Lotus japonicas (Lo). Number refers to amino acid residues.
Figure 3. The expression of OsRab7 in jasmine rice tissues. The expression of OsRab7 in various rice
tissues; roots = R, the third leaf blades at 20 and 50 DAG = Ll and L4, flower = F, and seeds at 15 DAF = S
(A). The relative expression level of OsRab7 in the rice tissues, presented as the ratio of expression of
OsRab7/β-Actin (B). β-Actin was used as an internal control.
114 Int. Res. J. Plant Sci.
Figure 4. The expression of OsRab7 under stress conditions. The expression of OsRab7 in rice leaves incubated
in 3 mM MES buffer pH 5.8 supplemented with 100 and 200 mM NaCl under continuous light for 3 and 5 days (A);
supplemented with 100 µM of ABA for 0, 3, 6, 12, and 24 h (B); supplemented with 1 mM ethephon for 0, 3, 6, 12,
and 24 h (C); or in leaves were kept in darkness for 0, 3 and 5 days (D); and wounded leaves at 0, 3 and 5 days
after wounding (E). The relative expression level of OsRab7 in those rice tissues, presented as the ratio of
expression of OsRab7/β-Actin (F-J). β-Actin was used as an internal control. The meaning of letter code; D = day,
h = hour.
Pitakrattananukool et al. 115
Figure 5. Chlorophyll content of rice leaf samples incubated under stress induced senescence conditions. The
chlorophyll content of the rice leaf samples incubated in 3 mM MES buffer pH 5.8 supplemented with 100 and 200
mM NaCl under continuous light for 0, 3 and 5 days (A); supplemented with 100 µM of ABA for 0, 3, 6, 12, and 24 h
(B); supplemented with 1 mM Ethephon for 0, 3, 6, 12, and 24 h (C); or in leaves kept in darkness for 0, 3 and 5
days (D); and wounded leaves at 0, 3 and 5 days after wounding (E). The meaning of letter the code: D = day, h =
hour.
upregulated in most of the stress induced-senescence
conditions except ethylene, its expression was
determined in leaves under natural senescence
conditions. The QPCR results revealed that the
expression levels of OsRab7 showed the highest
expression in leaf blades at 40 and 50 DAG [Figure 5(A)].
The relative level of expression was also analyzed from
the bands intensities [Figure 5(B)]. This shows that the
OsRab7 mRNA levels increased during leaf senescence
periods.
In this study OsRab7, was isolated from Thai jasmine
rice and examined with an expression analysis. The
result shows that the amino acids sequence retained the
Rab specific domains (Figure 2). Interestingly, OsRab7
showed the highest expression levels approximately 3fold up-regulated in senescing leaves over young leaves
and upregulated in leaves that were incubated in most
stress induced-senescence conditions. Moreover, the
decrease in chlorophyll content under the stress inducedsenescence was strong evidence that when leaves were
senescing, the expression of OsRab7 was high. In
addition, the OsRab7 sequence showed high similarity
(97%) to OsRab7B3 isolated from japonica rice which is
known to enhance leaf senescence in transgenic rice
(Pitakrattananukool et al., 2012). Therefore, OsRab7 is a
senescence-related gene likely to be involved in the
chlorophyll degradation process during leaf senescence
in Thai jasmine rice. For further analysis, OsRab7 gene
116 Int. Res. J. Plant Sci.
Figure 6. The expression of OsRab7 in jasmine rice leaves under natural senescence conditions. The expression
of OsRab7 in the third leaf blades at 20, 30, 40 and 50 DAG = Ll, L2, L3, and L4 respectively (A). The relative
expression levels of OsRab7 in those rice tissues, presented as the ratio of expression of OsRab/β-Actin (B). βActin was used as an internal control.
transfer is required to further understand its function in
the senescence process.
ACKNOWLEDGMENTS
This work was granted by the Office of the Higher
Education Commission, Thailand; Ms. Sugunya
Pitakrattananukool was supported by a CHE PhD.
Scholarship.
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