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R Zardoya

Publications and source records attributed to R Zardoya.

At least 19 recordsLinked to original sources

Differential population structuring of two closely related fish species, the mackerel (Scomber scombrus) and the chub mackerel (Scomber japonicus), in the Mediterranean Sea.

Population genetic structures of the mackerel (Scomber scombrus) and chub mackerel (Scomber japonicus) were studied in the Mediterranean Sea. Fragments of 272 bp (S. scomber) and 387 bp (S. japonicus) of the 5'-end of the mitochondrial control region were sequenced from spawning individuals collected off the coasts of Greece, Italy, Spain, and Portugal. High levels of mitochondrial control region haplotypic diversity (> 0.98) were found for both Scomber species. Nucleotide diversity was higher in the mackerel (0.022) than in the chub mackerel (0.017). Global F(ST) values were also higher and significant in the mackerel (0.024, P < 0.0001) as opposed to the chub mackerel (0.003, P > 0.05). Molecular variance analyses showed differential genetic structuring for these two closely related species. There is extensive gene flow between Mediterranean Sea and Atlantic Ocean populations of chub mackerel, which are organized into a larger panmictic unit. In contrast, Mediterranean Sea populations of mackerel show some degree of genetic differentiation and are structured along an east-west axis. The analysed eastern Mediterranean Sea mackerel populations (Greece, Italy) are clearly separated from that of the western Mediterranean Sea (Barcelona), which forms a panmictic unit with eastern Atlantic Ocean populations. The genetic structures of both species showed asymmetric migration patterns and indicated population expansion.

Analysis of Variance↗

On the origin of and phylogenetic relationships among living amphibians.

The phylogenetic relationships among the three orders of modern amphibians (Caudata, Gymnophiona, and Anura) have been estimated based on both morphological and molecular evidence. Most morphological and paleontological studies of living and fossil amphibians support the hypothesis that salamanders and frogs are sister lineages (the Batrachia hypothesis) and that caecilians are more distantly related. Previous interpretations of molecular data based on nuclear and mitochondrial rRNA sequences suggested that salamanders and caecilians are sister groups to the exclusion of frogs. In an attempt to resolve this apparent conflict, the complete mitochondrial genomes of a salamander (Mertensiella luschani) and a caecilian (Typhlonectes natans) were determined (16,656 and 17,005 bp, respectively) and compared with previously published sequences from a frog (Xenopus laevis) and several other groups of vertebrates. Phylogenetic analyses of the mitochondrial data supported with high bootstrap values the monophyly of living amphibians with respect to other living groups of tetrapods, and a sister group relationship of salamanders and frogs. The lack of phylogenetically informative sites in the previous rRNA data sets (because of its shorter size and higher among-site rate variation) likely explains the discrepancy between our results and those based on previous molecular data. Strong support of the Batrachia hypothesis from both molecule- and morphology-based studies provides a robust phylogenetic framework that will be helpful to comparative studies among the three living orders of amphibians and will permit better understanding of the considerably divergent vertebral, brain, and digit developmental patterns found in frogs and salamanders.

Amphibians↗

The evolutionary position of turtles revised.

Consensus on the evolutionary position of turtles within the amniote phylogeny has eluded evolutionary biologists for more than a century. This phylogenetic problem has remained unsolved partly because turtles have such a unique morphology that only few characters can be used to link them with any other group of amniotes. Among the many alternative hypotheses that have been postulated to explain the origin and phylogenetic relationships of turtles, a general agreement among paleontologists emerged in favoring the placement of turtles as the only living survivors of the anapsid reptiles (those that lack temporal fenestrae in the skull). However, recent morphological and molecular studies have radically changed our view of amniote phylogenetic relationships, and evidence is accumulating that supports the diapsid affinities of turtles. Molecular studies favor archosaurs (crocodiles and birds) as the living sister group of turtles, whereas morphological studies support lepidosaurs (tuatara, lizards, and snakes) as the closest living relatives of turtles. Accepting these hypotheses implies that turtles cannot be viewed any longer as primitive reptiles, and that they might have lost the temporal holes in the skull secondarily rather than never having had them.

Alligators and Crocodiles↗

A phylogenetic framework for the aquaporin family in eukaryotes.

A comprehensive evolutionary analysis of aquaporins, a family of intrinsic membrane proteins that function as water channels, was conducted to establish groups of homology (i.e., to identify orthologues and paralogues) within the family and to gain insights into the functional constraints acting on the structure of the aquaporin molecule structure. Aquaporins are present in all living organisms, and therefore, they provide an excellent opportunity to further our understanding of the broader biological significance of molecular evolution by gene duplication followed by functional and structural specialization. Based on the resulting phylogeny, the 153 channel proteins analyzed were classified into six major paralogous groups: (1) GLPs, or glycerol-transporting channel proteins, which include mammalian AQP3, AQP7, and AQP9, several nematode paralogues, a yeast paralogue, and Escherichia coli GLP; (2) AQPs, or aquaporins, which include metazoan AQP0, AQP1, AQP2, AQP4, AQP5, and AQP6; (3) PIPs, or plasma membrane intrinsic proteins of plants, which include PIP1 and PIP2; (4) TIPs, or tonoplast intrinsic proteins of plants, which include alphaTIP, gammaTIP, and deltaTIP; (5) NODs, or nodulins of plants; and (6) AQP8s, or metazoan aquaporin 8 proteins. Of these groups, AQPs, PIPs, and TIPs cluster together. According to the results, the capacity to transport glycerol shown by several members of the family was acquired only early in the history of the family. The new phylogeny reveals that several water channel proteins are misclassified and require reassignment, whereas several previously undetermined ones can now be classified with confidence. The deduced phylogenetic framework was used to characterize the molecular features of water channel proteins. Three motifs are common to all family members: AEF (Ala-Glu-Phe), which is located in the N-terminal domain; and two NPA (Asp-Pro-Ala) boxes, which are located in the center and C-terminal domains, respectively. Other residues are found to be conserved within the major groups but not among them. Overall, the PIP subfamily showed the least variation. In general, no radical amino acid replacements affecting tertiary structure were identified, with the exception of Ala-->Ser in the TIP subfamily. Constancy of rates of evolution was demonstrated within the different paralogues but rejected among several of them (GLP and NOD).

Amino Acid Motifs↗

Mitochondrial evidence on the phylogenetic position of caecilians (Amphibia: Gymnophiona).

The complete nucleotide sequence (17,005 bp) of the mitochondrial genome of the caecilian Typhlonectes natans (Gymnophiona, Amphibia) was determined. This molecule is characterized by two distinctive genomic features: there are seven large 109-bp tandem repeats in the control region, and the sequence for the putative origin of replication of the L strand can potentially fold into two alternative secondary structures (one including part of the tRNA(Cys)). The new sequence data were used to assess the phylogenetic position of caecilians and to gain insights into the origin of living amphibians (frogs, salamanders, and caecilians). Phylogenetic analyses of two data sets-one combining protein-coding genes and the other combining tRNA genes-strongly supported a caecilian + frog clade and, hence, monophyly of modern amphibians. These two data sets could not further resolve relationships among the coelacanth, lungfishes, and tetrapods, but strongly supported diapsid affinities of turtles. Phylogenetic relationships among a larger set of species of frogs, salamanders, and caecilians were estimated with a mitochondrial rRNA data set. Maximum parsimony analysis of this latter data set also recovered monophyly of living amphibians and favored a frog + salamander (Batrachia) relationship. However, bootstrap support was only moderate at these nodes. This is likely due to an extensive among-site rate heterogeneity in the rRNA data set and the narrow window of time in which the three main groups of living amphibians were originated.

Animals↗

Phylogenetic relationships of Greek cyprinidae: molecular evidence for at least two origins of the Greek cyprinid fauna.

A phylogeny of the main cyprinid species that live in Greece was based on the complete mitochondrial cytochrome b gene (1140 bp). The great diversity of the Greek cyprinid fauna, with at least 4 different lineages of barbins, 1 of gobionins, and 10 of leuciscinids, was confirmed. The validity and composition of the genera Tropidophoxinellus and Pachychilon were demonstrated. Likewise, the close phylogenetic relationships of genera Vimba, Blicca, and Abramis, as well as Telestes, Chondrostoma, and Phoxinellus, were established. Moreover, Pseudophoxinus beoticus was found to be closely related to Phoxinellus rather than to other Pseudophoxinus species. The most interesting outcome of our analyses was the existence of at least two different origins of the Greek cyprinid fauna. Most of the Greek cyprinids were directly related to the Danubian (i.e., central European) ichthyofauna. However, we found at least four cases of a close phylogenetic relationship between Greek and Iberian endemic species.

Animals↗

Molecular evidence on the evolutionary and biogeographical patterns of European cyprinids.

The phylogenetic relationships of 106 European cyprinid taxa were determined based on the complete nucleotide sequence (1140 bp) of the mitochondrial cytochrome b gene. The molecular phylogeny was used (1) to revise the current systematics of European cyprinids, (2) to establish the phylogenetic utility of traditional morphological characters that are widely used in Cyprinidae systematics, and (3) to discuss alternative hypotheses on the biogeography of the family in Europe. The age of the major lineages within European cyprinids was tentatively estimated with a molecular clock and showed full agreement with the fossil record of the group. Moreover, the results provided unambiguous evidence for a close phylogenetic affinity of some Caucasian and Greek endemic cyprinid taxa (e.g., B. capito and B. brachycephalus and Leuciscus keadicus, Barbus graecus, and B. albanicus, respectively) to Iberian and North African, but not Central European, cyprinids. The existence of such unexpected phylogenetic relationships refutes the classical hypothesis on the biogeography of European cyprinids, which assumes a dispersal of the cyprinid fauna from central Europe to southern Europe and northern Africa during the Miocene (and, hence, predicts a close phylogenetic relationship of all Caucasian, Greek, Iberian, and North African cyprinids to central European taxa). Instead, the existence of a Mediterranean realm independent of the central European route seems plausible based on the molecular evidence. It is likely that the new biogeographical scenario proposed here might apply to other primary freshwater European animals with low dispersal abilities, including fish, amphibians, and invertebrates.

Animals↗

Complete mitochondrial genome suggests diapsid affinities of turtles.

Despite more than a century of debate, the evolutionary position of turtles (Testudines) relative to other amniotes (reptiles, birds, and mammals) remains uncertain. One of the major impediments to resolving this important evolutionary problem is the highly distinctive and enigmatic morphology of turtles that led to their traditional placement apart from diapsid reptiles as sole descendants of presumably primitive anapsid reptiles. To address this question, the complete (16,787-bp) mitochondrial genome sequence of the African side-necked turtle (Pelomedusa subrufa) was determined. This molecule contains several unusual features: a (TA)n microsatellite in the control region, the absence of an origin of replication for the light strand in the WANCY region of five tRNA genes, an unusually long noncoding region separating the ND5 and ND6 genes, an overlap between ATPase 6 and COIII genes, and the existence of extra nucleotides in ND3 and ND4L putative ORFs. Phylogenetic analyses of the complete mitochondrial genome sequences supported the placement of turtles as the sister group of an alligator and chicken (Archosauria) clade. This result clearly rejects the Haematothermia hypothesis (a sister-group relationship between mammals and birds), as well as rejecting the placement of turtles as the most basal living amniotes. Moreover, evidence from both complete mitochondrial rRNA genes supports a sister-group relationship of turtles to Archosauria to the exclusion of Lepidosauria (tuatara, snakes, and lizards). These results challenge the classic view of turtles as the only survivors of primary anapsid reptiles and imply that turtles might have secondarily lost their skull fenestration.

Animals↗

Cloning and characterization of a microsatellite in the mitochondrial control region of the African side-necked turtle, Pelomedusa subrufa.

The nucleotide sequence of the African side-necked turtle mitochondrial control region and its flanking tRNA genes was determined. This 73% A+T-rich region is 1194 bp long. Several conserved motifs involved in the regulation of the mitochondrial genome replication process, including one conserved sequence block (CSB1), and three termination-associated sequences were identified. The most remarkable feature found in this control region was the presence of six microsatellite-containing tandem repeats between the CSB1 motif and the tRNAPhe gene. The potential usefulness of this microsatellite sequence for population-level studies is enhanced by its unique localization in the maternally inherited mitochondrial molecule.

Animals↗

Phylogenetic relationships of Iberian cyprinids: systematic and biogeographical implications.

The phylogenetic relationships among all Iberian endemic cyprinids were inferred using the complete nucleotide sequence of the cytochrome b gene. The inferred molecular phylogeny included representatives from Central European, Asian and North African species, and is highly congruent with previous phylogenies based on osteological characters. Iberian cyprinids were grouped into only five, very speciose lineages (with the exception of the monotypic Anaecypris): Barbus, Luciobarbus, Chondrostoma, Leuciscus and Anaecypris. The existence of such a relatively small number of Iberian cyprinid lineages can be explained by the historical isolation of the Iberian Peninsula. North African and Asian barbels are the sister group of Iberian Luciobarbus, supporting a south-eastern route of colonization of the Iberian Peninsula for this subgenus. With leuciscins, Anaecypris hispanica was considered a relict species as it could not be related to any other Iberian cyprinid. The phylogenetic relationships among the main lineages of Iberian cyprinids based on cytochrome b sequence data supported the traditional division of the Cyprinidae into two subfamilies: Cyprininae and Leuciscinae.

Animals↗

Platyrrhine systematics: a simultaneous analysis of molecular and morphological data.

Platyrrhine phylogeny has been investigated repeatedly with morphological characters and DNA nuclear gene sequences, with partially inconsistent results. Given the finding in the past decade that the mitochondrial genome is a potentially valuable source of phylogenetic information, we gathered DNA sequence data of a fragment of the 16S and the entire 12S mitochondrial genes. The objectives were to generate a cladistic phylogeny based on these data and to combine them in a simultaneous analysis with morphological characters and preexisting nuclear DNA sequences. Mitochondrial data analyzed on its own yielded a cladogram that was different from those generated with other data sets. The simultaneous analysis of mitochondrial, nuclear, and morphological data yielded a tree most congruent with that generated with nuclear data and to a lesser degree with the morphological one. It depicted a basal dichotomy that led to two major clades: one of them comprised [Atelinae (Callicebus + Pitheciini)] and the other major clade comprised [Aotus ((Cebus, Saimiri) (Callitrichinae))]. The weakest point of the phylogeny was the position of Aotus as basal within their clade as opposed to more closely linked with either the callitrichines or Cebus-Saimiri. Relationships within callitrichines and atelines were unstable as well. The simultaneous phylogenetic analysis of all data sets revealed congruent signal in all of them that was partially obscured in the separate analyses.

Amino Acid Sequence↗

Limitations of metazoan 18S rRNA sequence data: implications for reconstructing a phylogeny of the animal kingdom and inferring the reality of the Cambrian explosion.

We document the phylogenetic behavior of the 18S rRNA molecule in 67 taxa from 28 metazoan phyla and assess the effects of among-site rate variation on reconstructing phylogenies of the animal kingdom. This empirical assessment was undertaken to clarify further the limits of resolution of the 18S rRNA gene as a phylogenetic marker and to address the question of whether 18S rRNA phylogenies can be used as a source of evidence to infer the reality of a Cambrian explosion. A notable degree of among-site rate variation exists between different regions of the 18S rRNA molecule, as well as within all classes of secondary structure. There is a significant negative correlation between inferred number of nucleotide substitutions and phylogenetic information, as well as with the degree of substitutional saturation within the molecule. Base compositional differences both within and between taxa exist and, in certain lineages, may be associated with long branches and phylogenetic position. Importantly, excluding sites with different degrees of nucleotide substitution significantly influences the topology and degree of resolution of maximum-parsimony phylogenies as well as neighbor-joining phylogenies (corrected and uncorrected for among-site rate variation) reconstructed at the metazoan scale. Together, these data indicate that the 18S rRNA molecule is an unsuitable candidate for reconstructing the evolutionary history of all metazoan phyla, and that the polytomies, i.e., unresolved nodes within 18S rRNA phylogenies, cannot be used as a single or reliable source of evidence to support the hypothesis of a Cambrian explosion.

Animals↗

Searching for the closest living relative(s) of tetrapods through evolutionary analyses of mitochondrial and nuclear data.

The phylogenetic relationships of the African lungfish (Protopterus dolloi) and the coelacanth (Latimeria chalumnae) with respect to tetrapods were analyzed using complete mitochondrial genome DNA sequences. A lungfish + coelancanth clade was favored by maximum parsimony (although this result is dependent on which transition:transversion weights are applied), and a lungfish + tetrapod clade was supported by neighbor-joining and maximum-likelihood analyses. These two hypotheses received the strongest statistical and bootstrap support to the exclusion of the third alternative, the coelacanth + tetrapod sister group relationship. All mitochondrial protein coding genes combined favor a lungfish + tetrapod grouping. We can confidently reject the hypothesis that the coelacanth is the closest living relative of tetrapods. When the complete mitochondrial sequence data were combined with nuclear 28S rRNA gene data, a lungfish + coelacanth clade was supported by maximum parsimony and maximum likelihood, but a lungfish + tetrapod clade was favored by neighbor-joining. The seeming conflicting results based on different data sets and phylogenetic methods were typically not statistically strongly supported based on Kishino-Hasegawa and Templeton tests, although they were often supported by strong bootstrap values. Differences in rate of evolution of the different mitochondrial genes (slowly evolving genes such as the cytochrome oxidase and tRNA genes favored a lungfish + coelacanth clade, whereas genes of relatively faster substitution rate, such as several NADH dehydrogenase genes, supported a lungfish + tetrapod grouping), as well as the rapid radiation of the lineages back in the Devonian, rather than base compositional biases among taxa seem to be directly responsible for the remaining uncertainty in accepting one of the two alternate hypotheses.

Animals↗

Multilocus phylogeny of cichlid fishes (Pisces: Perciformes): evolutionary comparison of microsatellite and single-copy nuclear loci.

Among vertebrates, cichlid fishes are the paradigmatic example of adaptive radiation and ecological specialization. In turn, molecular genetic studies of cichlids have focused primarily on more recently diverged groups. Here, we present an evolutionary hypothesis of the major lineages of cichlid fishes based on DNA sequence data from two nuclear loci. One marker, Tmo-4C4, is a single-copy locus containing a region of amino acid similarity to the muscle protein TITIN. Flanking sequence from a second, microsatellite, locus Tmo-M27, shows similarity to mammalian RAS guanine nucleotide-releasing factor. We compare and combine data from these loci to evaluate phylogenetic performance. In separate and combined analyses, the sequence data support and clarify previous morphological hypotheses of cichlid major-group relationships. Indian and Malagasy cichlids form a basal, paraphyletic group. Neotropical cichlids are the sister clade to an African assemblage composed of the paraphyletic west and Pan-African lineages and a group of east African rift lake taxa. We use a consensus phylogeny of the Cichlidae to trace evolutionary changes in the microsatellite repeat motif at Tmo-M27. Analysis reveals that the repeat region was nearly lost in the ancestor to cichlids and then amplified extensively in African taxa. Results demonstrate that the two new DNA markers could be widely applied in perciform systematics. Furthermore, the comparative approach can unveil mutational dynamics of simple-sequence repeat loci over long periods of fish evolution. Simple-sequence repeat regions are increasingly being found in introns of important regulatory genes. We address issues involving their function and suggest caution in making assumptions of strict neutrality.

Animals↗

Molecular phylogenetic information on the identity of the closest living relative(s) of land vertebrates.

The phylogenetic position of tetrapods relative to the other two living sarcopterygian lineages (lungfishes and the coelacanth) has been subject to debate for many decades, yet remains unresolved. There are three possible alternatives for the phylogenetic relationships among these three living lineages of sarcopterygians, i.e., lungfish as living sister group of tetrapods, the coelacanth as closest living relative of tetrapods, and lungfish and coelacanth equally closely related to tetrapods. To resolve this important evolutionary question several molecular data sets have been collected in recent years, the largest being the almost complete 28S rRNA gene sequences (about 3500 bp) and the complete mitochondrial genomes of the coelacanth and a lungfish (about 16,500 bp each). Phylogenetic analyses of several molecular data sets had not provided unequivocal support for any of the three hypotheses. However, a lungfish + tetrapod or a lungfish + coelacanth clade were predominantly favored over a coelacanth + tetrapod grouping when the entire mitochondrial genomes alone or in combination with the nuclear 28S rRNA gene data were analyzed with maximum parsimony, neighbor-joining, and maximum likelihood phylogenetic methods. Also, current paleontological and morphological data seem to concur with these molecular results. Therefore the currently available molecular data seems to rule out a coelacanth + tetrapod relationship, the traditional textbook hypothesis. These tentative molecular phylogenetic results point to the inherent difficulty in resolving relationships among lineages which apparently originated in rapid succession during the Devonian.

Animals↗

The complete DNA sequence of the mitochondrial genome of a "living fossil," the coelacanth (Latimeria chalumnae).

The complete nucleotide sequence of the 16,407-bp mitochondrial genome of the coelacanth (Latimeria chalumnae) was determined. The coelacanth mitochondrial genome order is identical to the consensus vertebrate gene order which is also found in all ray-finned fishes, the lungfish, and most tetrapods. Base composition and codon usage also conform to typical vertebrate patterns. The entire mitochondrial genome was PCR-amplified with 24 sets of primers that are expected to amplify homologous regions in other related vertebrate species. Analyses of the control region of the coelacanth mitochondrial genome revealed the existence of four 22-bp tandem repeats close to its 3' end. The phylogenetic analyses of a large data set combining genes coding for rRNAs, tRNAs, and proteins (16,140 characters) confirmed the phylogenetic position of the coelacanth as a lobe-finned fish; it is more closely related to tetrapods than to ray-finned fishes. However, different phylogenetic methods applied to this largest available molecular data set were unable to resolve unambiguously the relationship of the coelacanth to the two other groups of extant lobe-finned fishes, the lungfishes and the tetrapods. Maximum parsimony favored a lungfish/coelacanth or a lungfish/tetrapod sistergroup relationship depending on which transversion:transition weighting is assumed. Neighbor-joining and maximum likelihood supported a lungfish/tetrapod sistergroup relationship.

Amino Acid Sequence↗

Evolutionary conservation of microsatellite flanking regions and their use in resolving the phylogeny of cichlid fishes (Pisces: Perciformes).

A phylogeny of the principal lineages of cichlid fishes and two other fish families of the suborder Labroidei was based on phylogenetic information from DNA sequences of the flanking region of a (CA)n microsatellite locus. Microsatellite (CA)n containing clones from a genomic library of an African cichlid fish from Lake Tanganyika, Tropheus moorii, were sequenced and primers for the polymerase chain reaction designed. All primers amplified the homologous microsatellite loci in many more than the source species and one microsatellite flanking locus (TmoM27) was particularly conserved and amplified in several lineages of perciform fishes that diverged more than 80-100 million years ago. Despite the extensive level of evolutionary conservation of this microsatellite flanking region (MFR), this nuclear region contained reliable phylogenetic information in the form of both point and length mutations. A phylogeny of cichlids based on this MFR agrees with other phylogenetic hypotheses based on morphological, mitochondrial, and anonymous nuclear DNA. Madagascan and Indian cichlids are found to be paraphyletic and the most basal group in the family Cichlidae. African and Neotropical cichlids are both monophyletic and sistergroups. Within African lineages, the East African cichlids are most likely to be monophyletic and the West African cichlids are probably paraphyletic and basal to all African species. The focal microsatellite locus contained much variation in (CA)n repeats in African cichlids and in surfperches (up to 64 repeats), but was short (with only 2-4 repeats) and almost invariant in Neotropical cichlids. The design of phylogenetically highly versatile MFR-primers will be of use not only for phylogeny reconstruction among families of perciform fishes, but also for population-level work in the thousands of species belonging to this highly species-rich suborder of fishes.

Animals↗

Evolutionary analyses of hedgehog and Hoxd-10 genes in fish species closely related to the zebrafish.

The study of development has relied primarily on the isolation of mutations in genes with specific functions in development and on the comparison of their expression patterns in normal and mutant phenotypes. Comparative evolutionary analyses can complement these approaches. Phylogenetic analyses of Sonic hedgehog (Shh) and Hoxd-10 genes from 18 cyprinid fish species closely related to the zebrafish provide novel insights into the functional constraints acting on Shh. Our results confirm and extend those gained from expression and crystalline structure analyses of this gene. Unexpectedly, exon 1 of Shh is found to be almost invariant even in third codon positions among these morphologically divergent species suggesting that this exon encodes for a functionally important domain of the hedgehog protein. This is surprising because the main functional domain of Shh had been thought to be that encoded by exon 2. Comparisons of Shh and Hoxd-10 gene sequences and of resulting gene trees document higher evolutionary constraints on the former than on the latter. This might be indicative of more general evolutionary patterns in networks of developmental regulatory genes interacting in a hierarchical fashion. The presence of four members of the hedgehog gene family in cyprinid fishes was documented and their homologies to known hedgehog genes in other vertebrates were established.

Amino Acid Sequence↗