Supplementary materials
Figure S1. The maximum-likelihood tree of OsCASP1 homologs with
their protein sequences. The scale bar indicates the simple matching
distance. The alignment and phylogenetic tree were constructed with
MEGAX
(http://megasoftware.net/). The
numbers for interior branches indicate the bootstrap values for 500
replications.
Figure S2 . (A) Alignment of CASPs in Arabidopsis and
rice. IC: Intracellular loop; TM: Transmembrane domain; EC:
extracellular loop. (B) (C) Transmembrane domains of OsCASP1 was
predicted with TMHMM 2.0 (http://www.cbs.dtu.dk/services/TMHMM/) (B) and
Protter
(http://wlab.ethz.ch/protter/#)
(C)
Figure S3. Models of the constructs for complementation test,
tissue-specific expression, and subcellular localization of OsCASP1.CASP1-N includes a transmembrane region predicted by TMHMM with a lower
probability. TM: Transmembrane region.
Figure S4. The phenotypes of the WT and els1 mutant and
characterization of els1.(A) Internode length and plant height; IN = internode. (B) The tiller
number of the WT and els1 mutant at the heading stage. (C) and
(D) The tiller phenotype of the seedling at 33-d-old. The emergence of
tiller in the els1 mutant was delayed. (E) The els1 locus
was fine-mapped on chromosome 4. The numbers of recombinants are shown
in brackets.
Figure S5. The genotype and phenotype of CRISPR/cas9 mutants.(A) Schematic map of OsCASP1 gene and sequence alignment of the
sgRNA target region showing altered bases in different mutant lines. The
target region of the sgRNA was indicate with arrow, and the heterozygous
transgenic plants with different mutant sites were marked with
rectangles. (B) The typical phenotypes of CRISPR/cas9 transgenic plants.
Figure S6. Subcellular localizations of OsCASP1 protein. (A)OsCASP1pro:OsCASP1-GFP in rice root. (B) and (C)OsCASP1pro:OsCASP1-GFP in onion and Arabidopsisprotoplast. (D) and (E) 35S::DsRed-OsCASP1 in onion andArabidopsis protoplast. Circle indicates the nuclear position.
(F) 35S::DsRed-CASP1-N in rice protoplasts. CASP1-N includes a
transmembrane region predicted by TMHMM with a lower probability. ER:
Endoplasmic reticulum.
Figure S7. Subcellular localizations of OsCASP1 protein in
transgenic plant roots carrying 35S::DsRed-OsCASP1. (A) OsCASP1
in the cell membrane in small lateral roots. (B) and (C) OsCASP1 is
concentrated in the stele of lateral roots. (D) and (E) OsCASP1 in main
roots. Arrows indicate the nuclei.
Figure S8. The localization of OsCASP1pro:OsCASP1-GUSexpression after salt treatment. (A) Whole roots treated with NaCl. (B)
Crown root with lateral roots. (C) Lateral root. (D) Transection of the
crown root. (E) Longitudinal section of the crown root. ep, epidermis;
ex, exodermis; sc, sclerenchyma; co, cortex; en, endodermis; pe,
pericycle. Scale bar: (A) 1cm; (B), (C), (D) and (E) 0.1cm.
Figure S9. The representative cross-sections at different zones
in primary roots stained with berberine-aniline blue and
phloroglucinol. (A) Staining with berberine-aniline blue.
Autofluorescence of cell walls is detected as blue. Arrow head indicates
CS at endodermis. (B) Staining with phloroglucinol. Arrow head indicates
endodermis. ep, epidermis; ex, exodermis; sc, sclerenchyma.
Figure S10. The representative cross-sections stained with
periodic acid (A) and permeability of CS was evaluated using periodic
acid (B). Purple color indicates that the periodic acid penetrated into
root tissues. (C) The roots stained with periodic acid.