Electrophoretic characterization of Amaranthus L. seed proteins and ...

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60 R. JUAN ET AL.67674330433020.120.114.414.41 2 3 4 5 6 7 8 9 10 11Figure 1. Tricine-sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) of Amaranthus seed proteins.1, A. viridis (V-3, V-2, V-1); 2, A. powellii ssp. bouchonii (P-1); 3, A. muricatus (M-2, M-1, M-3); 4, A. deflexus (D-3, D-2,D-1); 5, A. graecizans ssp. sylvestris (G-1); 6, A. blitoides (B-3, B-2, B-1); 7, A. retroflexus (R-2, R-3, R-1); 8, A. blitum (Bl-2,Bl-1); 9, A. albus (A-2, A-3, A-1); 10, A. cruentus (C-3, C-2, C-1); 11, A. hypochondriacus (H-1, H-2).The SDS-PAGE profile of the Amaranthus seed proteinsstudied showed a range of peptides varying from64 to 12 kDa (Fig. 1). The most abundant peptideswere in the ranges 64–49.2 kDa and 36.8–32.8 kDa,although the largest number of protein bands wereobserved between 25.1 and 12 kDa. Fourteen bandswere considered for the taxonomic study, althoughsome were subdivided into several isoforms based onthe differences observed in Rf.A similarity analysis based on the SDS-PAGEprofile of seed proteins was carried out. The softwareused displayed a single tree from all those possible(Fig. 2). Two major clusters (A and B) with about 45%similarity were obtained. The first cluster (group A)includes A. powellii ssp. bouchonii, A. graecizans ssp.sylvestris, and A. retroflexus, and the remaining taxastudied form group B. Within group B, two subgroups(C and D) were distinguished with 58% similarity.Group C includes five species, with A. deflexus as themost dissimilar taxon (63%). The other taxa of thisgroup form two well-defined clusters: A. muricatusand A. viridis with 79% similarity, and A. blitoidesand A. blitum with 72% similarity. With respect to thelast group (group D), A. albus clearly differs fromA. cruentus and A. hypochondriacus (72%), whosepopulations appear to be partly mixed.By contrast with the complexity of the identificationof Amaranthus species using morphological characters,our results showed that the Amaranthus taxastudied are well defined based on the seed proteinprofile; in particular, species such as A. blitoidesshowed a similarity higher than 95% in the populationsstudied. Thus, except for A. hypochondriacusand A. cruentus, the UPGMA dendrogram arrangesthe studied species in single clusters, indicating thatthe data provided by the electrophoresis profile ofseed proteins are useful in the discrimination of Amaranthustaxa at the specific level. The fact that thepopulations of A. cruentus and A. hypochondriacusare mixed could be the result of two different mechanisms.Firstly, a hybridization phenomenon, which isvery frequent in these species (Costea, Sanders &Waines, 2001a). In this case, population C3 ofA. cruentus could be a hybrid population betweenA. cruentus and A. hypochondriacus. This is possibleas this population shares the same peptide band kDawith A. hypochondriacus populations, which was notobserved in the other A. cruentus populations.Sammour (1991) also observed an intermediateprotein profile between A. viridis and A. hybridus(subgenus Albersia) in a population of A. viridis, indicatinga hybridization process between the two taxa.The second explanation for the protein profile of thispopulation could be the hybrid origin of A. hypochondriacusfrom A. cruentus and A. powellii, as proposedby Sauer (1993) and supported by some molecularstudies (Transue et al., 1994; Chan & Sun, 1997).Although the dendrogram obtained from theseed protein profile (Fig. 2) discriminates betweenspecies, it does not group these species according tothe infrageneric classification shown in Table 1.Thus, relationships established by the electrophoreticprofile of seed proteins do not match theestablished relationships based on morphological ortraditional characters. For example, A. blitum andA. blitoides, morphologically different and assigned© 2007 The Linnean Society of London, Botanical Journal of the Linnean Society, 2007, 155, 57–63
SEED PROTEIN PROFILE OF AMARANTHUS 61Figure 2. Clustering based on the seed protein profiles of 11 taxa of Amaranthus according to the Bray–Curtis similarityindex (1 - dissimilarity index ¥ 100). V-1, V-2, V-3, A. viridis; P-1, A. powellii ssp. bouchonii; M-2, M-1, M-3, A. muricatus;D-3, D-2, D-1, A. deflexus; G-1, A. graecizans ssp. sylvestris; B-3, B-2, B-1, A. blitoides; R-2, R-3, R-1, A. retroflexus; Bl-2,Bl-1, A. blitum; A-2, A-3, A-1, A. albus; C-3, C-2, C-1, A. cruentus; H-1, H-2, A. hypochondriacus.to different sections (Mosyakin & Robertson, 1996),show a similar protein profile, indicating a closerrelationship. In addition, A. albus, A. cruentus, andA. hypochondriacus are in the same group (D),although they are assigned to different sections andeven different subgenera: A. albus belongs to subgenusAlbersia section Pyxidium, whereas A. cruentusand A. hypochondriacus are included in subgenusAmaranthus section Amaranthus. Although, from amorphological point of view, A. albus is very differentfrom A. cruentus and A. hypochondriacus,recently, Costea & DeMason (2001) found similaritybetween them using leaf anatomical features, asthey observed intermediate characters between bothsubgenera in A. albus. The fact that species belongingto subgenus Albersia are distributed betweenthe three main groups (A, C, D) is congruent withtheir heterogeneous taxa composition, and supportsthe opinion of Mosyakin & Robertson (1996) thatthis subgenus needs a taxonomic revision in order todelimit new sections.In agreement with Sammour (1991) and Zheleznovet al. (1997), a correlation between the electrophoresisprofile of seed proteins and the karyological data isobserved in Amaranthus. Thus, included in group Aare taxa (A. powellii ssp. bouchonii, A. graecizans ssp.sylvestris, and A. retroflexus) with basic numbersx = 16 and 17 (Pastor, 1992). In group D, all taxa showx = 16, whereas, in group C, all species show x = 17,except for A. blitoides, which has x = 16 (Pastor,1992). This could mean that the seed protein profile isone of the phenotypical differences determined by thebasic number.In conclusion, seed proteins in Amaranthus areuseful characters to discriminate between species. Inaddition, these characters show low environmentaland evolutionary variability. The profile obtained bySDS-PAGE of seed proteins provides interestinginformation that will doubtless help in the clarificationof this complex genus. Our results show theclose relationships between species belonging todifferent taxonomic groups, such as sections orsubgenera, and allow a hybrid population to be distinguished.A more detailed study is necessary inorder to clarify the infrageneric classification of thisgenus.© 2007 The Linnean Society of London, Botanical Journal of the Linnean Society, 2007, 155, 57–63
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