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10 Supplemental Table 1. The progeny of crosses between MPK4/mpk4-2 and ANQ/anq-2. Genotype Number Number Number observed expected-1 expected-2 (1) MPK4/MPK4 ANQ/ANQ (2) MPK4/MPK4 ANQ/anq (3) MPK4/MPK4 anq-2/anq (4) MPK4/mpk4-2 ANQ/ANQ (5) MPK4/mpk4-2 ANQ/anq (6) MPK4/mpk4-2 anq-2/anq (7) mpk4-2/mpk4-2 ANQ/ANQ (8) mpk4-2/mpk4-2 ANQ/anq (9) mpk4-2/mpk4-2 anq-2/anq Total number P-value 1.5E-12<0.05* 0.46>0.05 The genotypes of 197 progeny plants with the respective indicated genotypes were determined by PCR. Expected values were calculated using Mendel s law (expected-1). Expected values were also calculated by reference to the viability of the female gametophyte and/or the embryo, as summarized in Table 2 (expected-2). P-values were calculated by the χ 2 -test from observed and expected values. A P- value indicative of a significant difference is
11 indicated by an asterisk.
12 Supplemental Table 2. The frequency of cells with an incomplete cell plate. Genotype Number of Number of The frequency of cells with an cells in cells with an incomplete cell one incomplete cell plate plate in one section (b) (%) section (a) Col-0 # Col-0 # Col-0 # Col-0 # Col-0 # Col-0 # Average frequency 0 ± 0 mpk13 # mpk13 # mpk13 # mpk13 # mpk13 # mpk13 # Average frequency 0 ± 0 mpk4-2 #
13 mpk4-2 # mpk4-2 # mpk4-2 # mpk4-2 # mpk4-2 # Average frequency 2.5 ± 0.62 P-value * 2.0E-06<0.05 mpk4-2 mpk13 # mpk4-2 mpk13 # mpk4-2 mpk13 # mpk4-2 mpk13 # mpk4-2 mpk13 # mpk4-2 mpk13 # Average frequency 2.1 ± 0.61 P-value * 7.0E-06<0.05 P-value ** 0.37>0.05 (a-b) Transverse sections (5 µm) were prepared from cotyledons of 9-day-old Col-0, mpk13, mpk4-2, and mpk4-2 mpk13 seedlings. The number of cells and the number of cells with incomplete cell plates were counted in a typical single section from cotyledons of six homozygous siblings (#1 to #6). Average frequencies are presented as means ± SD (n = 6). P-values were calculated by Student s t-test. A single asterisk indicates comparison with Col-0. Two asterisks indicate comparison with mpk4-2.
14 Supplemental Table 3. Primers used in this study. Application Sequence (5-3 ) (forward/reverse) Genotyping of MPK4 ATTAAGCTTCTCAAGCATATGGATCATGA/ TCCTCCTTGAAATATCTACAGAGTTGGTGT Genotyping of mpk4-1 mutant CCGGATCGTATCGGTTTTCG /CTTGAGTCATCAGGAGATCC Genotyping of mpk4-2 mutant GCGTGGACCGCTTGCTGCAACT/ TCCTCCTTGAAATATCTACAGAGTTGGTGT Genotyping of ANQ/MKK6 CTACGCGTCTTGAGATCTTG/ GGATTGAGAGAAAGCGTTAAGATCAGAG Genotyping of anq-2 mutant GCGTGGACCGCTTGCTGCAACT/ GGATTGAGAGAAAGCGTTAAGATCAGAG Genotyping of MPK13 GAAGATGGAGGGATCTTGAC/ CAGACAAGCTCCTTGATGTC Genotyping of mpk13 mutant GAAGATGGAGGGATCTTGAC/ CATTTTATAATAACGCTGCGGACATCTAC
15 Genotyping of MPK11 CACACTAAGAATCTTAAGGGTTAAACTTGACT G/ATGTCAATAGAGAAACCATTCTTCGGTG Genotyping of mpk11 mutant GCGTGGACCGCTTGCTGCAACT /ATGTCAATAGAGAAACCATTCTTCGGTG Genotyping of NACK1 GCGGCCATATACCTTACAGA/ GCTCTCCGATTTCCATTTCCA Genotyping of nack1-3 mutant GCGGCCATATACCTTACAGA/ GCGTGGACCGCTTGCTGCAACT RT-PCR analysis of MPK4 TGCTCTGAATACACAGCAGC/ CACACTGAGTCTTGAGGATTG Quantitative RT-PCR analysis β-6-tubulin ACCACTCCTAGCTTTGGTGATCTG/ AGGTTCACTGCGAGCTTCCTCA Quantitative RT-PCR analysis EF1-a TGAGCACGCTCTTCTTGCTTTCA/ GGTGGTGGCATCCATCTTGTTACA Quantitative RT-PCR analysis of MPK4 CACAGTTGATGAGGCGTTGTG/ CACATACCGGTTCTTCGTTGATAT Quantitative RT-PCR analysis of MPK5 ACGGTTGAGGAAGCACTGTGT/ GAACAAACAGGCTCGTCGTTAA
16 Quantitative RT-PCR analysis of MPK11 TTTGACCCAAACAGACGCATT/ TCGTGTAGCGGTGCTAAGTAAGG Quantitative RT-PCR analysis of MPK12 CCTAACCGGCGCATCTCA/ TCATGGTGCGGTGATAGGTAAG Quantitative RT-PCR analysis of MPK13 K4_5p_F GCATTGAAGCAGCCGTACTTG/ CAAAGCTGAAAGGAGTAGGACATG GGTACCGACTTGTTTGTGAATATAGAGGAAA CATG (Underline = Kpn1 site) K4_5p_R GGATCCCACTGAGTCTTGAGGATTGAACTTGA C (Underline = BamH1 site) K4_3p_F GGATCCGAGAAAGAGAGAGAGATATATATCC (Underline = BamH1 site) K4_3p_R GCGGCCGCGATAATTAGTGGATGTAATTAGA GTTAAGAC YFP_F GGATCCATGGTGAGCAAGGGCGAGGAGCTG (Underline = BamH1 site) K4WT_S K4WT_R LB6316 RT-PCR analysis of MPK11 GTGACAATGCAAGAAGATACGTTAGACAGC CTTGAAATATCTACAGAGTTGGTGTG TCAAACAGGATTTTCGCCTGCT AACCGAGTAACTTGCTCCTG CACAAGGCTTCATCGACTGT
17 KK 93 GCGGCCGCGACACTGAGTCTTGAGGATTGAA KK 138 GTCGACGATCTCGAGGCTGTTGAAAC (Underline = Sal1 site) Cloning of MPK4 CCATGGCGGCGGAGAGTTGTTTCG GCGGCCGCCACTGAGTCTTGAGGATTGAA Cloning of MPK5 CCATGGCGAAGGAAATTGAATCA GCGGCCGCAATGCTCGGCAGAGGATTGAA Cloning of MPK11 CCATGGCAATAGAGAAACCATTC GCGGCCGCAGGGTTAAACTTGACTGATTC Cloning of MPK12 CCATGGCTGGAGAATCAAGCTCT GCGGCCGCGTGGTCAGGATTGAATTTGACAGA Cloning of MPK13 CCATGGAGAAAAGGGAAGATGGA GCGGCCGCCATATTCTTGAAGTGTAAAGACTCT
18 Cloning of MPK1 CCATGGCGACTTTGGTTGATCC GCGGCCGCGAGCTCAGTGTTTAAGGTTG Cloning of MPK6 CCATGGACGGTGGTTCAGGTC GCGGCCGCTTGCTGATATTCTGGATTGAAA Cloning of MKK6/ANQ CCATGGTGAAGATCAAATCGAA GCGGCCGCTCTAAGGTAGTTAACAGGTG Cloning of MPK2 CCATGGCGACTCCTGTTGATCC GCGGCCGCAAACTCAGAGACCTCATTGT Cloning of MPK3 CCATGGACACCGGCGGTGGCCA GCGGCCGCACCGTATGTTGGATTGAGTG Cloning of MPK7 CCATGGCGATGTTAGTTGAGCC GCGGCCGCGGCATTTGAGATTTCAGCTT
19 Cloning of MPK8 CCATGGGTGGTGGTGGGAATCT GCGGCCGCAGAATTGTGAAGAGAAGCAA Cloning of MPK14 CCATGGCGATGCTAGTTGATCC GCGGCCGCAGCTCGGGGGAGGTAATGAA Cloning of MPK15 CCATGGGTGGTGGTGGCAATCT GCGGCCGCAGAATTGTGTAGAGATGCAA Cloning of MPK16 CCATGGAGCCTGATCACCGCAA GCGGCCGCATACCAGCGACTCATTGCAG Cloning of MPK17 CCATGGTGGAGAAAGAGTTTTT GCGGCCGCTGACACTGCAGAGGAGACAC Cloning of MPK18 CCATGGAACAAAATCAAGTGAA GCGGCCGCTGATGCTGCGCTGTAACTAA
20 Cloning of MPK19 CCATGGAAAAAACTCAGGAGAA GCGGCCGCAGACATGCCATACCCAACAG Supplemental Methods RT-PCR We used 15-day-old seedlings for analyses by RT-PCR, which was performed as described in Methods. Primers used for the experiments are listed in Supplemental Table 3. Plant materials and growth conditions Tobacco suspension-cultured BY-2 cells were maintained as described elsewhere (Banno et al., 1993; Nishihama et al., 2001). Transformed BY-2 cells were generated by Agrobacterium-mediated transformation (An. 1985). Fluorescence microscopy To generate the MPK4-GFP fusions, MPK4-coding regions without a termination codon were cloned into the pgwb5 vector. To generate the GFP-MAP65-3 fusions, MAP65-3-coding regions were cloned into the pgwb6 vector (Nakagawa et al. 2001). Living BY-2 cells that expressed MPK4-GFP were stained with 0.5 µg/ml Hoechst (Dojin, Kumamoto, Japan). Aniline blue (0.1% in 0.1 M K 3 PO 4, ph 11) was added to roots of 4-day-old plants on a Gellan gum plate and imaged after a 5-min incubation.
21 All techniques for microscopic visualization and analysis were described previously (Nishihama et al., 2001; Tanaka et al., 2001). Production and purification of recombinant proteins His-T7-tagged-MPK4, GST-fused MKK6/ANQ, and GST-fused-MAP65-3/PLE proteins were produced in E. coli BL21 using pet28 (Novagen) and pgex (GE Healthcare) vectors. His-T7-MPK4 was purified with Ni Sepharose TM 6 Fast Flow (GE Healthcare). GST-NQK1 and GST-MAP65-3 proteins were purified with Glutathione-Sepharose TM 4B (GE Healthcare). Phosphorylation assays in vitro His-T7-MPK4 and GST-MKK6/ANQ were incubated in 200 µl of kinase buffer (as described in Methods) that contained 500 µm ATP at 25 C for 120 min. Reaction products, which included His-T7-MPK4 (150 ng) and GST-MKK6/ANQ (225 ng), were incubated with GST-MAP65-3-PLE (1 mg) in 20 µl of kinase buffer that contained 50 µm ATP and 10 µci of [γ- 32 P] ATP (GE Healthcare) at 25 C for 60 min. Reactions were terminated by the addition of 10 ml of 3 SDS sample buffer and the mixtures were boiled. The samples were subjected to SDS-PAGE and radioactivity was visualized with an imaging analyzer (BAS-1800; Fujifilm, Tokyo, Japan).
22 Supplemental References An, G. (1985). High Efficiency Transformation of Cultured Tobacco Cells. Plant Physiol. 79: Banno, H., Hirano, K., Nakamura, T., Irie, K., Nomoto, S., Matsumoto, K., and Machida, Y. (1993). NPK1, a tobacco gene that encodes a protein with a domain homologous to yeast BCK1, STE11, and Byr2 protein kinases. Mol. Cell. Biol. 13: Nakagawa, T., Kurose, T., Hino, T., Tanaka, K., Kawamukai, M., Niwa, Y., Toyooka, K., Matsuoka, K., Jinbo, T., and Kimura, T. (2007). Development of series of gateway binary vectors, pgwbs, for realizing efficient construction of fusion genes for plant transformation. J. Biosci. Bioeng. 104: Tanaka, H., Onouchi, H., Kondo, M., Hara-Nishimura, I., Nishimura, M., Machida, C., and Machida, Y. (2001). A subtilisin-like serine protease is required for epidermal surface formation in Arabidopsis embryos and juvenile plants. Development 128: Nishihama, R., Ishikawa, M., Araki, S., Soyano, T., Asada, T., and Machida, Y. (2001). The NPK1 mitogen-activated protein kinase kinase kinase is a regulator of cell-plate formation in plant cytokinesis. Genes Dev. 15:
At E17.5, the embryos were rinsed in phosphate-buffered saline (PBS) and immersed in
Supplementary Materials and Methods Barrier function assays At E17.5, the embryos were rinsed in phosphate-buffered saline (PBS) and immersed in acidic X-gal mix (100 mm phosphate buffer at ph4.3, 3 mm
Supplemental Data. Lee et al. Plant Cell. (2010) /tpc Supplemental Figure 1. Protein and Gene Structures of DWA1 and DWA2.
Supplemental Figure 1. Protein and Gene Structures of DWA1 and DWA2. (A) Protein structures of DWA1 and DWA2. WD40 region was determined based on the NCBI conserved domain databases (B, C) Schematic representation
Fertilization Recovery after Defective. Sperm Cell Release in Arabidopsis. Current Biology, Volume 22 Supplemental Information
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www.sciencesignaling.org/cgi/content/full/5/244/ra72/dc1 Supplementary Materials for An Interaction Between BZR1 and DELLAs Mediates Direct Signaling Crosstalk Between Brassinosteroids and Gibberellins
Supplemental Data. Liu et al. (2013). Plant Cell /tpc
Supplemental Figure 1. The GFP Tag Does Not Disturb the Physiological Functions of WDL3. (A) RT-PCR analysis of WDL3 expression in wild-type, WDL3-GFP, and WDL3 (without the GFP tag) transgenic seedlings.
Supplemental Data. Furlan et al. Plant Cell (2017) /tpc
Supplemental Data. Furlan et al. Plant Cell (0) 0.0/tpc..00. Supplemental Data. Furlan et al. Plant Cell (0) 0.0/tpc..00. Supplemental Data. Furlan et al. Plant Cell (0) 0.0/tpc..00. Supplemental Figure.
Supplementary Fig 1. The responses of ERF109 to different hormones and stresses. (a to k) The induced expression of ERF109 in 7-day-old Arabidopsis
Supplementary Fig 1. The responses of ERF109 to different hormones and stresses. (a to k) The induced expression of ERF109 in 7-day-old Arabidopsis seedlings expressing ERF109pro-GUS. The GUS staining
Supplementary information
Supplementary information The E3 ligase RNF8 regulates KU80 removal and NHEJ repair Lin Feng 1, Junjie Chen 1 1 Department of Experimental Radiation Oncology, The University of Texas M. D. Anderson Cancer
Supplemental Figure 1. Conserved regions of the kinase domain of PEPR1, PEPR2, CLV1 and BRI1.
Supplemental Figure 1. Conserved regions of the kinase domain of PEPR1, PEPR2, CLV1 and BRI1. The asterisks and colons indicate the important residues for ATP-binding pocket and substrate binding pocket,
Supplemental Data. Dai et al. (2013). Plant Cell /tpc Absolute FyPP3. Absolute
A FyPP1 Absolute B FyPP3 Absolute Dry seeds Imbibed 24 hours Dry seeds Imbibed 24 hours C ABI5 Absolute Dry seeds Imbibed 24 hours Supplemental Figure 1. Expression of FyPP1, FyPP3 and ABI5 during seed
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Supplemental Data Supplemental Figure 1. MYC2 Binds to the E-box but not the E1-box of the MPK6 Promoter. (A) E1-box and E-box (wild type) containing MPK6 promoter fragment. The region shown in red denotes
S156AT168AY175A (AAA) were purified as GST-fusion proteins and incubated with GSTfused
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 Supplemental Materials Supplemental Figure S1 (a) Phenotype of the wild type and grik1-2 grik2-1 plants after 8 days in darkness.
Supplementary Figures 1-12
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PCR Cloning II fusion domains for purification His 6 GST chitin binding protein MBP
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- 1 - Supplemental Data
- 1-1 Supplemental Data 2 3 4 5 6 7 8 9 Supplemental Figure S1. Differential expression of AtPIP Genes in DC3000-inoculated plants. Gene expression in leaves was analyzed by real-time RT-PCR and expression
Data Sheet Quick PCR Cloning Kit
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PIE1 ARP6 SWC6 KU70 ARP6 PIE1. HSA SNF2_N HELICc SANT. pie1-3 A1,A2 K1,K2 K1,K3 K3,LB2 A3, A4 A3,LB1 A1,A2 K1,K2 K1,K3. swc6-1 A3,A4.
A B N-terminal SWC2 H2A.Z SWC6 ARP6 PIE1 HSA SNF2_N HELICc SANT C pie1-3 D PIE1 ARP6 5 Kb A1 200 bp A3 A2 LB1 arp6-3 A4 E A1,A2 A3, A4 A3,LB1 K1,K2 K1,K3 K3,LB2 SWC6 swc6-1 A1,A2 A3,A4 K1,K2 K1,K3 100
(phosphatase tensin) domain is shown in dark gray, the FH1 domain in black, and the
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Supplemental Information
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pt7ht vector and over-expressed in E. coli as inclusion bodies. Cells were lysed in 6 M
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Supplementary information
Supplementary information Supplementary figures Figure S1 Level of mycdet1 protein in DET1 OE-1, OE-2 and OE-3 transgenic lines. Total protein extract from wild type Col0, det1-1 mutant and DET1 OE lines
SUPPLEMENTAL DATA. Supplementary data
1 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 SUPPLEMENTAL DATA Supplementary data Figure S1: Verification of pla-iα1 knockdown lines a: Semiquantitative
Figure S1. Figure S2. Figure S3 HB Anti-FSP27 (COOH-terminal peptide) Ab. Anti-GST-FSP27(45-127) Ab.
/ 36B4 mrna ratio Figure S1 * 2. 1.6 1.2.8 *.4 control TNFα BRL49653 Figure S2 Su bw AT p iw Anti- (COOH-terminal peptide) Ab Blot : Anti-GST-(45-127) Ab β-actin Figure S3 HB2 HW AT BA T Figure S4 A TAG
Construction of plant complementation vector and generation of transgenic plants
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Product Name : Competent Cell BL21(DE3)pLysS Code No. : DS260 Size : 100 μl 10 Competency : > 5 10 7 cfu/μg (puc19) Supplied product : SOC medium, 1 ml 10 This product is for research use only Description
Supplemental Data. Shimoda et al. (2011). Plant Cell /tpc B C D E
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Supplemental Figure 1
Supplemental Figure 1 A gta2-1 gta2-2 1kb AT4G08350 B Col-0 gta2-1 LP+RP+LBb1.3 C Col-0 gta2-2 LP+RP+LB1 D Col-0 gta2-2 gta2-1 GTA2 TUBLIN E Col0 gta2-1 gta2-2 Supplemental Figure 1. Phenotypic analysis
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Supplemental Figure 1 HDA18 has an HDAC domain and therefore has concentration dependent and TSA inhibited histone deacetylase activity.
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Supporting Information
Supporting Information Üstün and Börnke, 2015 Figure S1: XopJ does not display acetyltransferase activity. Autoacetylation activity in vitro. Acetylation reactions using MBP, MBP-XopJ, GST and GST-HopZ1a
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Local auxin metabolism regulates environment-induced hypocotyl elongation Zuyu Zheng 1,2, Yongxia Guo 3, Ondřej Novák 4,5, William Chen 2, Karin Ljung 4, Joseph P. Noel 1,3, *, and Joanne Chory 1,2, *
indicated numbers of pups at day of life (DOL) 10, or embryonic day (ED) B. Male mice of
SUPPLEMENTRY FIGURE LEGENDS Figure S1. USP44 loss leads to chromosome missegregation.. Genotypes obtained from the indicated numbers of pups at day of life (DOL) 10, or embryonic day (ED) 13.5.. Male mice
Supplemental Data. Aung et al. (2011). Plant Cell /tpc
35S pro:pmd1-yfp 10 µm Supplemental Figure 1. -terminal YFP fusion of PMD1 (PMD1-YFP, in green) is localized to the cytosol. grobacterium cells harboring 35S pro :PMD1-YFP were infiltrated into tobacco
Supplemental Data. Seo et al. (2014). Plant Cell /tpc
Supplemental Figure 1. Protein alignment of ABD1 from other model organisms. The alignment was performed with H. sapiens DCAF8, M. musculus DCAF8 and O. sativa Os10g0544500. The WD40 domains are underlined.
Supplemental Data. Guo et al. (2015). Plant Cell /tpc
Supplemental Figure 1. The Mutant exb1-d Displayed Pleiotropic Phenotypes and Produced Branches in the Axils of Cotyledons. (A) Branches were developed in exb1-d but not in wild-type plants. (B) and (C)
Expanded View Figures
Expanded View Figures Figure EV1. AM3-CLE45 control experiments and bam3 alleles. A Relative primary root length of indicated genotypes at 9 dag, in response to increasing amounts of CLE45 in the media.
pri 201 DNA Series (High-Expression Vectors for Plant Transformation)
For Research Use Cat #. 3264 3265 3266 3267 pri 201 DNA Series (High-Expression Vectors for Plant Transformation) Product Manual Table of Contents I. Description... 3 II. Product Information... 3 III.
SUPPLEMENTAL INFORMATION
SUPPLEMENTAL INFORMATION SUPPLEMENTAL DATA Table S1: Related to Figure 4. DnaA screen for divalent cations and nucleotides. Concentration of reagents, T m and F fold30 75. Reagent Conc T m (±SEM) mm C
Supplemental Data. Farmer et al. (2010) Plant Cell /tpc
Supplemental Figure 1. Amino acid sequence comparison of RAD23 proteins. Identical and similar residues are shown in the black and gray boxes, respectively. Dots denote gaps. The sequence of plant Ub is
GTTCGGGTTCC TTTTGAGCAG
Supplementary Figures Splice variants of the SIP1 transcripts play a role in nodule organogenesis in Lotus japonicus. Wang C, Zhu H, Jin L, Chen T, Wang L, Kang H, Hong Z, Zhang Z. 5 UTR CDS 3 UTR TCTCAACCATCCTTTGTCTGCTTCCGCCGCATGGGTGAGGTCATTTTGTCTAGATGACGTGCAATTTACAATGA
Supplementary Table 1. The Q-PCR primer sequence is summarized in the following table.
Supplementary Table 1. The Q-PCR primer sequence is summarized in the following table. Name Sequence (5-3 ) Application Flag-u ggactacaaggacgacgatgac Shared upstream primer for all the amplifications of
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