figure s1. schematic diagram of vector pu1391w used to study subcellular localization of proteins....
TRANSCRIPT
![Page 1: Figure S1. Schematic diagram of vector pU1391W used to study subcellular localization of proteins. RB and LB, right and left T-DNA border;; Hpt, hygromycin](https://reader036.vdocument.in/reader036/viewer/2022083009/5697c02a1a28abf838cd820b/html5/thumbnails/1.jpg)
Figure S1. Schematic diagram of vector pU1391W used to study subcellular localization of proteins. RB and LB, right and left T-DNA border;; Hpt, hygromycin phosphotransferase gene; GFP, green fluorescent protein gene; PWRKY13, rice OsWRKY13 promoter; NOS, napoline synthase polyadenylation signal.
RB PWRKY13 target gene GFP NOS Hpt LB
HindIII PstI BamHI EcoRI
![Page 2: Figure S1. Schematic diagram of vector pU1391W used to study subcellular localization of proteins. RB and LB, right and left T-DNA border;; Hpt, hygromycin](https://reader036.vdocument.in/reader036/viewer/2022083009/5697c02a1a28abf838cd820b/html5/thumbnails/2.jpg)
Figure S2. Gel mobility shift assays. The electrophoresis was performed with the gel plates of 6 cm in length, which could not clearly distinguish different sizes of DNA-protein complexes.
The nuclear proteins were from 8 h after mock-inoculated (control) or JL691-inoculated Mudanjiang 8. DNA probes are 39- to 51-bp in length (Supplemental Table 2).
(a) D
53I8
D53
I10
D53
I12
D53
I14
D53
I16
D53
I18
D53
I20
D53
I22
D53
I24
Mock inoculation
D53
I8D
53I1
0
D53
I12
D53
I14
D53
I16
D53
I18
D53
I20
D53
I22
D53
I24
Pathogen (JL691) inoculation
![Page 3: Figure S1. Schematic diagram of vector pU1391W used to study subcellular localization of proteins. RB and LB, right and left T-DNA border;; Hpt, hygromycin](https://reader036.vdocument.in/reader036/viewer/2022083009/5697c02a1a28abf838cd820b/html5/thumbnails/3.jpg)
D53
I12-
1 (p
ath
ogen
inoc
ula
tion
)
D53
I12-
2 (p
ath
ogen
inoc
ula
tion
)
D53
I16-
1 (p
ath
ogen
inoc
ula
tion
)
D53
I16-
2 (p
ath
ogen
inoc
ula
tion
)
To determine the pathogen-responsive DNA-protein binding regions in probes D53I12 and D53I16 as shown in Supplemental Fig. 2a, four 17- to 25-bp probes (Supplemental Table 2) from the two long probes were examined for their binding ability to nuclear proteins from pathogen-inoculated rice cultivar Mudanjiang 8. The gel mobility shift assay shows that D53I8-1, D53I12-1 do not contain protein-binding site and D53I12-2, D53I16-1 and D53I16-2 contain protein-binding sites.
(b)
![Page 4: Figure S1. Schematic diagram of vector pU1391W used to study subcellular localization of proteins. RB and LB, right and left T-DNA border;; Hpt, hygromycin](https://reader036.vdocument.in/reader036/viewer/2022083009/5697c02a1a28abf838cd820b/html5/thumbnails/4.jpg)
To further define the pathogen-responsive DNA-protein binding regions in probes D53I12-2 and D54I16-2 as shown in Supplemental Fig. 2b, four 9- to 13-bp probes (Supplementary Table 2) from the two long probes were examined for their binding ability to nuclear proteins from pathogen-inoculated rice cultivar Mudanjiang 8. The gel mobility shift assay shows that D53I12-2-1, D53I12-2-2 and D53I16-2-2 contain protein-binding sites and D53I16-2-1 do not contain protein-binding site.
D53
I12-
2-1
(pat
hog
en in
ocu
lati
on)
D53
I12-
2-2
(pat
hog
en in
ocu
lati
on)
D53
I16-
2-1
(pat
hog
en in
ocu
lati
on)
D53
I16-
2-2
(pat
hog
en in
ocu
lati
on)
(c)
![Page 5: Figure S1. Schematic diagram of vector pU1391W used to study subcellular localization of proteins. RB and LB, right and left T-DNA border;; Hpt, hygromycin](https://reader036.vdocument.in/reader036/viewer/2022083009/5697c02a1a28abf838cd820b/html5/thumbnails/5.jpg)
To further define the pathogen-responsive DNA-protein binding regions in probes D53I12-2-1and D53I16-2-2 as shown in Supplemental Fig. 2c, six 5- to 7-bp probes (Supplementary Table 2) from the two long probes were examined for their binding ability to nuclear proteins from pathogen-inoculated rice cultivar Mudanjiang 8. The gel mobility shift assays show that all the probes contain protein-binding sites, indicating that the protein-binding sites in the two probes shown in Supplemental Fig. 1C can not be further defined.
(d)
D53
I12-
2-1-
1 (p
ath
ogen
inoc
ula
tion
)
D53
I12-
2-1-
2 (p
ath
ogen
inoc
ula
tion
)
D53
I12-
2-1-
3 (p
ath
ogen
inoc
ula
tion
)
D53
I16-
2-2-
1 (p
ath
ogen
inoc
ula
tion
)
D53
I16-
2-2-
2 (p
ath
ogen
inoc
ula
tion
)
D53
I16-
2-2-
3 (p
ath
ogen
inoc
ula
tion
)
![Page 6: Figure S1. Schematic diagram of vector pU1391W used to study subcellular localization of proteins. RB and LB, right and left T-DNA border;; Hpt, hygromycin](https://reader036.vdocument.in/reader036/viewer/2022083009/5697c02a1a28abf838cd820b/html5/thumbnails/6.jpg)
(e)
D53
I16-
2-2
(M)
D53
I12-
2-1
(P)
D53
I16-
2-2
(P)
D53
I12-
2-1
(M)
The nuclear proteins were from 8 h after mock-inoculated (M, control) or pathogen (JL691)-inoculated (P) Mudanjiang 8. The gel mobility shift assay shows that D53I12-2-1 and D53I16-2-2 contain protein-binding sites, which have the same protein binding patterns as D53I12 and D54I16 shown in Supplemental Fig. 2a.