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Supplementary data:
Supplementary Tables
Table S1. Primers used to generate the expression vectors and restriction site sequences
underlined
Primer
s
Amplification of
the
5’ (F)
sites
GGCCTTAATTAACCCAGTCCTCGCG
native
Pac I
GTA
promoter of
5’ (R)
OsPT1
Amplification
Incorporated restriction
Sequences
TTATGGCGCGCCGGCTTCCCAACTC
Asc I
TTT
of
5’ (F)
GCACTAGTGCCGCAAAGCAAAAGA
ORF of OsPT1
Spe I
AAAAG
5’ (R)
CGACTAGTCGGAAGCGACGACTAG
Spe I
AGTT
Generation of the
5’ (F)
GCTGGGTACCACTAGTTTTCATGGC
OsPT1-RNAi
construct
Kpn I/Spe I
GATCGTCA
5’ (R)
GATAGGATCCGAGCTCATGAGCGTG
BamH I/Sac I
AATCCATAGAC
Table S2. Primers used to amplify the OsPT1 cDNA fragments
For RT-PCR analysis
For qRT-PCR analysis
Primers
Sequences
OsPT1 5’ (F)
AGGCACTCATCGCACTCT
OsPT1 5’ (R)
GCATACCGAGGAAGTTTGT
Actin 5’
(F)
GGAACTGGTATGGTCAAGGC
Actin 5’
(R)
AGTCTCATGGATAACCGCAG
OsPT1 5’ (F)
CGCTTCCGTACGAGTGGTAGT
OsPT1 5’ (R)
Actin 5’ (F)
GGTTCTTTCAAATCCAGGGAAA
TTATGGTTGGGATGGGACA
Actin 5’ (R)
AGCACGGCTTGAATAGCG
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Figure S1
Roots
–P
Leaves
+P
–P
+P
OsPT1
OsActin
Figure S1 The transcriptional patterns of OsPT1 in roots and leaves of rice (Oryza
sativa L.ssp. Japonica cv. Nipponbare).
10-d-old rice seedlings were transferred to the Pi-sufficient (300 µM Pi, +P) and
Pi-deficient (10 µM Pi, -P) conditions for 21d. RNA was extracted from roots and
leaves of rice plant and determined by semi-quantitative RT-PCR for OsPT1 (28
cycles) and actin (26 cycles). A housekeeping gene, actin (OsRac1, accession number
AB047313), was used as the internal standard.
Figure S2
Ubi -GUS
OsPT1
Figure S2 The expression of OsPT1 driven by the native promoter in reproductive
organs of rice.
OsPT1 promoter-driven expression of the GUS reporter gene in spikelets (left) and
generating buds (right) of the rice grown in the soil are shown. Ubiquitin
promoter-driven expression of the GUS reporter gene was used as positive control.
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Figure S3 Relative expression of the genes that encode Pi-transporters belonging to
Pht1 family in shoot of OsPT1 overexpression transgenic and wild type plants.
10-d-old rice seedlings were transferred to Pi-sufficient (300 µM Pi) solution for 21d.
Total RNAs were extracted from the shoots of the seedlings. Relative expression of
the Pi-transporters belonging to Pht1 family were determined by real time quantitative
RT-PCR. OsPT11 and OsPT13, AMF colonization enhanced PiTs, had not been
detected. A housekeeping gene, actin (OsRac1, accession number AB047313), was
used as the internal standard.
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Figure S4
Ox1 Ox2
Ri1
Ri2
WT
Figure S4 33Pi uptake in OsPT1-Ox, OsPT1-Ri and wild-type plants.
OsPT1-Ox, OsPT1-Ri and wild-type plants were grown for 7d and then transferred
into Pi-sufficient (300 µM Pi) medium for 3 d. The Pi uptake of these 10-d-old
seedlings was monitored over a 12h period.
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Figure S5
Figure S5 Motif analysis in the putative promoters of Pi-transporter genes belonging
to Pht1 family in rice.
The relative positions of the cis-regulatory elements identified by the RSA-Tools
(http://rsat.ulb.ac.be/rsat/) program are marked within each promoter sequence with
symbols specific for each type of motif: W-box (TTGACY) represented by a blue
square; PHO-like element (CDHGTGG) represented by a red dual-triangle; P1BS
(GNATATNC) represented by a green circle.
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