Down-regulation of SymRK correlates with a deficiency in vascular bundle development in Phaseolus vulgaris nodules
Version of Record online: 8 SEP 2011
© 2011 Blackwell Publishing Ltd
Plant, Cell & Environment
Volume 34, Issue 12, pages 2109–2121, December 2011
How to Cite
SÁNCHEZ-LÓPEZ, R., JÁUREGUI, D., NAVA, N., ALVARADO-AFFANTRANGER, X., MONTIEL, J., SANTANA, O., SANCHEZ, F. and QUINTO, C. (2011), Down-regulation of SymRK correlates with a deficiency in vascular bundle development in Phaseolus vulgaris nodules. Plant, Cell & Environment, 34: 2109–2121. doi: 10.1111/j.1365-3040.2011.02408.x
- Issue online: 9 NOV 2011
- Version of Record online: 8 SEP 2011
- Accepted manuscript online: 16 AUG 2011 09:45AM EST
- Received 6 June 2011; received in revised form 28 July 2011; accepted for publication 2 August 2011
Figure S1. (a) Alignment of predicted amino acid sequences of SymRK-like receptor orthologs. Sequences were aligned using ClustalX version 2.0.6. Accession numbers are as indicated in Materials and Methods. (b) Scheme of the primary structure of PvSymRK. Protein domains are indicated: signal peptide (SP), non-conserved extracellular (EC), extracellular leucine-rich regions (Leu), transmembrane domain (TM) and intracellular kinase-like domain (IC kinase). Positions of the PvSymRK primary structure fragments used to design the RNAi specific for PvSymRK (RNAi) are indicated. Position of the fragment amplified in the qRT-PCR analysis and the probe used in the Southern blot experiment are illustrated.
Figure S2. Genomic Southern blot analysis of PvSymRK sequences. Lanes correspond to Phaseolus vulgaris genomic DNA (10 µg) fully digested with EcoRI, BglII, EcoRV or HindIII, as indicated. Fragment used as probe was as indicated in Supporting Information Fig. S1. Size markers (MWM) are in kbp.
Figure S3. Specificity of polyclonal anti-PvSymRK antibodies, as determined by Western blot analysis. (A) Whole extracts from wild-type (Wild-type) roots or hairy roots expressing PvSymRK tagged with a cMyc epitope at the C-terminal (PvSymRK-cMyc) were incubated with either pre-immune serum, anti-PvSymRK (α-PvSymRK) or anti-cMyc E910 monoclonal antibody (α-cMyc), as indicated. (B) Whole extracts from nodules (15 dpi) incubated with anti-PvSymRK, as indicated. Molecular weight markers are in kDa.
Figure S4. Negative control in immunofluorescence experiments. No immunofluorescence signal has been detected using pre-immune serum. 15 dpi nodule section. Scale bar is as indicated.
Figure S5. Expression of two LRR-RLK receptors (NARK and KLV) in PvSymRKi hairy roots. Transcript accumulation in transgenic hairy roots (pTDT and PvSymRKi) was assessed by qRT-PCR using gene-specific primers (PvNARK-Up, 5′-GGTGGATCCAAGGCTCAGTGGG-3′; PvNARK-Lw, 5′-AGGAGGATTGGTGAGCATGTGAAC-3′; PvKLV-Up, 5′-GTGCTTCACTTGGCAATTGTGTGC-3′; PvKLV-Lw, 5′-AAGATGAAGCCACAGAAACGGGC-3′). PvNARK (GB: CV544058.1) and PvKLV (PhvGI: TC38989) sequences were obtained by in silico searching in available P. vulgaris sequence databases.
Figure S6. Histological analysis of nodule primordia induced in P. vulgaris wild-type roots (5 dpi). (A) Section containing two nodule primordia at different stages (5 dpi and ∼3 dpi) as well as a fragment of a lateral root primordia, as indicated. (B) Large magnification of the proximal zone of the nodule primordial (5 dpi) shown in A. Arrowheads indicate pro-vascular traces. Scale bars are as indicated.
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