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Table S1 (Ho et al.)
Table S1 Primers and their sequences used in this study.
Name
Sequence
CDPK1-F1
5’-CTACAGATCTATGGGGAATCAGTGCCA-3'
CDPK1-R1
5'-TCATAGATCTCTAAACTCCATTTTCACAGAT-3'
CDPK1-F2
5'-TCTAGGGTACCGGATCCGATCGAGCGCGCGAGG-3'
CDPK1-R2
5'-CTCAACTGCAGATCCAACCCCCGCCTGAG-3'
GFP-F3
5'-TCTAGCTGCAGGTGAGCAAGGGCGAG-3'
GFP-R3.1
5'-TCTAGGTCGACTTACTTGTACAGCTCGTC-3'
GFP-R3.2
5'-TCTAGGTCGACACGCTGAACTTGTGGC-3'
GF14c-F4
5'-TAAGGATCCATGTCTCGGGAGGAGAAT-3'
GF14c-R4
5'-TCAGGATCCTTACTGGCCCTCGCAGGCGT-3'
GF14c-F5
5'-TCTAGGGTACCGGATCCACGAGACCACTCGAACCCGACCCGCCTC-3'
GF14c-R5
5'-ACCTTAGATTCTGCAGCAGTAGATGAG-3'
CDPK1-F6
5’-ATGGGGAATCAGTGCCAGAAC-3’
CDPK1-R6
5’-AAGTTGTGCCAAACTGGCCCT-3’
Gene accession number: OsCDPK1 (AY158077), GFP (U43284), OsGF14c (U65957).
Restriction enzyme recognition sites are underlined.
.
Fig. S1 (Ho et al.)
1
MGNQCQNGTL GSDYHNRFPR EHAVGYVQGD SYLDLKKFDD TWPEVNNFKP
51
TAASILRRGL DPTSINVLGR KTADLREHYI IGRKLGQAQF GTTYLCTEIN
101
TGCEYACKTI PKRKLITKED VEDVRREIQI MHHLSGHKNV VAIKDVYEDG
151
QAVHIVMELC AGGELFDRIQ EKGHYSERKA AELIRIIVSI VAMCHSLGVM
P
VIb
VII
VIII
HRDLKPENFL LLDKDDDLSI KAIDFGLSVF FKPGQVFTEL VGSPYYVAPE
201
251
301
351
401
451
501
I
II
III
IV
VIa
V
IX
VLHKRYGPES DVWSAGVILY VLLSGVPPFW AETQQGIFDA VLKGHIDFQS
W
X
DPWPKISDSA KDLIRKMLSH CPSERLKAHE VLRHPWICEN GVATDQALDP
SVISRLKQFS AMNKLKKLAL RVIAERLSEE EIAGLREMFK AVDTKNRGVI
E
TFGELREGLR RFGAEFKDTE IGDIMEAAHN DNNVTIHYEE FIAATLPLNK
IEREEHLLAA FTYFDKDGSG YITVDKLQRA CGEHNMEDSL LEEIISEVDQ
R
Q
NNDGQIDYAE FVAMMQGSNV GLGWQTMESS LNVALRDAPQ VH*
R
P
Fig. S1
Structure and amino acids sequence of OsCDPK1.
Subdomains conserved in protein kinases are indicated by Roman numerals and
underlined. Four EF-hand structures of calcium binding regions are indicated by dash
underlines. Double dash underline indicates autoinhibitory regions of kinase activity.
Green and red letters represent changes of amino acid residue from OsCDPK1 to
OsCDPK12 and to OsCDPK13, respectively.
Fig. S2 (Ho et al.)
Fig. S2
Expression of OsCDPK1 in T4 transgenic rice. Total RNA was isolated from 14day-old seedlings and subjected to Northern blot analysis using the OsCDPK1specific probe. Ethidium bromide staining detected 25S and 18S rRNA bands. Lane
1, OsCDPK1 knockdown transgenic line (Ri-1); lane 2, OsCDPK1(tr)
overexpressing transgenic line (OEtr-1); lane 3, wild-type (Wt).
Fig. S3 (Ho et al.)
A
Wt
B
Ri1, 2, 3, 4, 5
OEtr1, 3, 4, 6, 7, 8, 9,15
OEtr-1
60 days
Wt
Ri-1
Fig. S3
Comparison of plant height between T1 generation of OsCDPK1(tr) (8 independent
lines) and OsCDPK1(Ri) (5 independent lines) and wild type (Wt) plants.
(A)The independent T1 transgenic plants were corresponding to the transformed cell
lines as showed in Figure 2, respectively, were grown in the isolated field for 60
days. (B) The T1 transgenic plants of OEtr-1 and Ri-1, and wild type plants were
planted in an another isolated field and grown for 60 days.
Fig. S4 (Ho et al.)
A
20.0
10 days
C
+GA3
20.0
10 days
16.0
12.0
12.0
Length (cm)
Length (cm)
10 days
16.0
8.0
8.0
4.0
4.0
0.0
0.0
Wt
OEtr-1
-GA3
Ri-1
Wt
OEtr-1
Ri-1
D
B
20.00
Seedlings length (cm)
Seedlins length (cm)
25.00
20.00
15.00
10.00
5.00
0.00
15.00
10.00
5.00
0.00
Wt
OEtr-1
Ri-1
Wt
OEtr-1
Ri-1
Fig. S4
The semi-dwarf phenotype of the OEtr-1 line was recovered by GA3 treatment.
Comparison of plant height between T4 generation of OEtr-1 and Ri-1 and wild
type (Wt) plants. Seeds were germinating on medium containing1/2 MS salt and
supplemented with (A and B) or without (C and D) 10 μM GA3 for 10 days.
Fig. S5 (Ho et al.)
Wt OEtr-1 Ri-1
OsCDPK1
GA20ox1
GA20ox2
GA3ox2
Act1
Fig. S5
The expression of GA biosynthesis-related genes in OEtr-1 and Ri-1 plants.
Total RNA was isolated from 14-day-old seedlings and subjected to RT-PCR
analysis using primers specific for OsCDPK1, GA20ox1, GA20ox2, and GA3ox2;
the rice Act1 gene was used as an internal control.
Fig. S6 (Ho et al.)
Wt
GF14c-Ox GF14c-Ri
Fig. S6
Drought tolerance in transgenic rice is enhanced by overexpression of the 14-3-3
protein GF14c.
Rice seedlings were grown on 1/2 MS agar medium for 14 days, transferred to
the pot and grown for another 7 days, withheld water for 5 days, and re-watered
for 4 days.