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Fig. 4 | BMC Evolutionary Biology

Fig. 4

From: Alternative translation initiation codons for the plastid maturase MatK: unraveling the pseudogene misconception in the Orchidaceae

Fig. 4

Immunoblot detection of MatK from orchid species. Orchid protein was resolved by SDS-PAGE and transferred to nitrocellulose membrane. MatK protein was detected using anti-MatK antibody as described in Barthet and Hilu [12]. Orchid species analysed represent two different subfamilies of Orchidaceae (Orchidoideae: Spiranthes vernalis, S. cernua, S. sinensis, Caladenia catenata and Cryptostylis erecta and Epidendroideae: Phaius tancarvilleae) and are representative of orchids that require the alternative initiation codon for full-length MatK translation (Figure 2A and B). All immunoblots were repeated twice to verify results. (a) Immunoblot detection of MatK from 50 μg of total protein from Spiranthes vernalis, S. cernua and Phaius tancarvilleae. N = 3 biological replicates. Mass standard = PageRuler Prestained Protein Ladder (Thermo Scientific). Ponceau S stain of RbcS shown as loading control. (b) Immunoblot detection of MatK from 75 μg of total protein from Caladenia catenata, Cryptostylis erecta and Spiranthes sinensis. N = 1 biological replicate due to tissue limitations. Oryza sativa (rice) was used as a control for detection. Mass standard = 6–185 kDa Protein Ladder (NEB). Ponceau S stain of RbcS shown as loading control. All immunoblots were repeated twice to verify results. (c) Alignment of MatK peptide region used for antibody generation to orchid species examined in this study

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