Patricia Iturra

The Atlantic salmon genome provides insights into rediploidization

The whole-genome duplication 80 million years ago of the common ancestor of salmonids (salmonid-specific fourth vertebrate whole-genome duplication, Ss4R) provides unique opportunities to learn about the evolutionary fate of a duplicated vertebrate genome in 70 extant lineages. Here we present a high-quality genome assembly for Atlantic salmon (Salmo salar), and show that large genomic reorganizations, coinciding with bursts of transposon-mediated repeat expansions, were crucial for the post-Ss4R rediploidization process. Comparisons of duplicate gene expression patterns across a wide range of tissues with orthologous genes from a pre-Ss4R outgroup unexpectedly demonstrate far more instances of neofunctionalization than subfunctionalization. Surprisingly, we find that genes that were retained as duplicates after the teleost-specific whole-genome duplication 320 million years ago were not more likely to be retained after the Ss4R, and that the duplicate retention was not influenced to a great extent by the nature of the predicted protein interactions of the gene products. Finally, we demonstrate that the Atlantic salmon assembly can serve as a reference sequence for the study of other salmonids for a range of purposes.

Sequencing the genome of the Atlantic salmon (Salmo salar)

The International Collaboration to Sequence the Atlantic Salmon Genome (ICSASG) will produce a genome sequence that identifies and physically maps all genes in the Atlantic salmon genome and acts as a reference sequence for other salmonids.

Identification of sex chromosome molecular markers using RAPDs and fluorescent in situ hybridization in rainbow trout

The goal of this work is to identify molecular markers associated with the sex chromosomes in rainbow trout to study the mode of sex determination mechanisms in this species. Using the RAPD assay and bulked segregant analysis, two markers were identified that generated polymorphic bands amplifying preferentially in males of the Mount Lassen and Scottish strains of rainbow trout. Chromosomal localization using fluorescent in situ hybridization of a 900 bp probe developed from one of these markers revealed a brightly defined signal on a chromosome that could morphologically be classified as the Y chromosome.

Phylogenetic relationships of Chilean leptodactylids: a molecular approach based on mitochondrial genes 12S and 16S

La mayoria de los anfibios chilenos pertenece a la subfamilia Telmatobiinae (Anura, Leptodactylidae). Varios estudios filogeneticos de Leptodactylidae y Telmatobiinae, basados principalmente en caracteres morfologicos, han sugerido implicitamente relaciones mas estrechas de algunas especies de Telmatobiinae con miembros de otras subfamilias de leptodactilidos, incluyendo el genero leptodactilino Pleurodema presente en Chile. Ademas, un numero creciente de estudios moleculares sugieren un estatus no monofiletico para Telmatobiinae, aunque ninguno de estos estudios ha investigado las relaciones filogeneticas de esta subfamilia. Secuencias parciales de los genes ribosomales mitocondriales 12S y 16S fueron comparadas para determinar las relaciones filogeneticas de los leptodactilidos chilenos y su posicion dentro de los anuros modernos (Neobatrachia). Se incluyeron 22 especies de nueve de los diez generos de telmatobinos presentes en Chile (Alsodes, Atelognathus, Batrachyla, Caudiverbera, Eupsophus, Hylorina, Insuetophrynus, Telmatobufo y Telmatobius), dos especies del genero Pleurodema y una especie de Rhinodermatidae la cual es considerada una familia derivada de los leptodactilidos por algunos autores. Se incluyeron ademas 51 especies que representan la mayoria de las familias que componen Neobatrachia. Las reconstrucciones filogeneticas se realizaron utilizando los metodos de maxima parsimonia, maxima verosimilitud e inferencia bayesiana. Las topologias obtenidas en todos los analisis indican que Telmatobiinae es un ensamblaje polifiletico, compuesto por especies que pertenecen a Hyloidea (la mayoria de los generos) y especies mas relacionadas con taxa de Australasia (el clado Caudiverbera + Telmatobufo, definido como la tribu Calyptocephalellini). Estos datos moleculares respaldan agrupaciones basadas en otro tipo de evidencia (Caudiverbera + Telmatobufo, Alsodes + Eupsophus y Batrachyla + Hylorina) y plantean nuevas hipotesis de relaciones para algunos generos de telmatobinos (Atelognathus con Batrachyla e Hylorina, Insuetophrynus + Rhinoderma). Las relaciones filogeneticas observadas en este estudio sugieren un origen multiple para los anuros del bosque templado de Chile y revelan un inesperado nivel de diversidad taxonomica y divergencia evolutiva entre los leptodactilidos chilenos

Characterization of sex chromosomes in rainbow trout and coho salmon using fluorescence in situ hybridization (FISH).

With the aim of characterizing the sex chromosomes of rainbow trout (Oncorhynchus mykiss) and to identify the sex chromosomes of coho salmon (O. kisutch), we used molecular markers OmyP9, 5S rDNA, and a growth hormone gene fragment (GH2), as FISH probes. Metaphase chromosomes were obtained from lymphocyte cultures from farm specimens of rainbow trout and coho salmon. Rainbow trout sex marker OmyP9 hybridizes on the sex chromosomes of rainbow trout, while in coho salmon, fluorescent signals were localized in the medial region of the long arm of one subtelocentric chromosome pair. This hybridization pattern together with the hybridization of a GH2 intron probe on a chromosome pair having the same morphology, suggests that a subtelocentric pair could be the sex chromosomes in this species. We confirm that in rainbow trout, one of the two loci for 5S rDNA genes is on the X chromosome. In males of this species that lack a heteromorphic sex pair (XX males), the 5S rDNA probe hybridized to both subtelocentrics This finding is discussed in relation to the hypothesis of intraspecific polymorphism of sex chromosomes in rainbow trout.

Diploid chromosome number variations and sex chromosome polymorphism in five cultured strains of rainbow trout (Oncorhynchus mykiss)

Abstract This work reveals the chromosomal characteristics of the five cultured strains of rainbow trout, Oncorhynchus mykiss. The five strains—Americana, Donaldson, Cofradex, Steelhead, and Kamloops—are those commonly used in the Chilean salmon farming industry. Chromosome characterization involved the determination of diploid chromosome number (2n), chromosome arm number (NF) and sex chromosome of individuals by Hoechst 33258/Actinomicyn D (H/AmD) fluorescent stain. Considering all of the strains studied, the 2n ranged from 58 to 63 chromosomes. All of the karyotypes had NF=104. Within this range of variation, the strains exhibited low (Americana, 2n=59–60; Steelhead and Kamloops, 2n=58–61), intermediate (Cofradex, 2n=59–62) or high chromosome numbers (Donaldson, 2n=61–63). In strains with the most extreme diploid chromosome numbers, the most frequent 2n corresponded to 59 and 61 chromosomes. Sex chromosome determination revealed that 24.5% of specimens of both sexes, presented an unexpected sex chromosome. In males, “XX” specimens were observed, whereas in females, “XY” or “XXX” individuals were recorded. The subtelocentric (st) morphology and centromeric H/AmD banding pattern of the additional “X” chromosome in “XX males” and “XXX females” is similar to that observed in the regular X chromosome. The chromosome number variation in the strains studied is consistent with the Robertsonian-type polymorphism described for rainbow trout. The presence of different chromosome numbers among strains, could reflect the chromosome differences existing within ancestral trout populations, or the presence of a certain degree of mixing with other populations of different geographical origin. The discovery of specimens with an unexpected sex chromosome is suggestive of an incomplete differentiation of the sex chromosome pair in the rainbow trout.

Physiological factors affecting triploid production in rainbow trout, Oncorhynchus mykiss

The influence of three physiological factors related to triploid production was examined for rainbow trout. Individual spawnings of 46 females were used to perform triploid production experiments from 1989 to 1991, with various water temperatures (6–13.8°C) at stripping and egg incubation. The percentage of triploids was similar for a heat shock of 26.5°C for 15 min applied 15 or 25 min after fertilization (73.4±5.6% vs 78.6±4.5%). A tendency to a higher percentage of triploids (85.9±5.7%) was observed for water temperatures of 6–8.0°C at stripping and incubation, compared to at higher water temperatures of 12.1–14°C (63.0±8.4%). Analysis of variance indicated an effect of four categories of water temperature (6–8, 8.1–10, 10.1–12, 12.1–14°C) on triploid product at P = 0.14. Regression analysis confirmed these as significant (r = −0.36; P < 0.02). A significant increase in percentage of triploids was observed when eggs remained 2, 6 and 10 days in the body cavity. The mean % triploidy was 46.1, 54.7 and 76.8, respectively. Similar results were observed for survival of the eggs.

Development and characterization of DNA sequence OmyP9 associated with the sex chromosomes in rainbow trout.

This work describes the construction and characterization of a sequence characterized amplified DNA region (SCAR DNA), designated OmyP9, that was derived from a RAPD marker associated with the sex chromosomes in rainbow trout. A RsaI restriction fragment length polymorphism in OmyP9 identifies variants A, B and C. We found six OmyP9 variant phenotypes – A, B, C, AB, BC and ABC, in 186 individuals of seven different rainbow trout strains. The patterns of inheritance of OmyP9 in 139 fingerlings from 10 crosses of three strains of rainbow trout were studied. The males had a greater representation of the A variant (93.3%) suggesting an association with the Y chromosome. All male fingerlings analysed inherited the A variant from their male parents. These results support the hypothesis that OmyP9 is located on the sex chromosomes of rainbow trout, and that for the males studied the A variant is located on the Y chromosome in a region close to sex determinants and/or in a sector where the genetic recombination between X and Y is restricted. The present evidence also supports our previous hypothesis that OmyP9 is organized as a tandem repeated sequence in the sex chromosomes of rainbow trout. We feel that the OmyP9 RsaI marker can be used for sex identification in crosses where it is possible to determine the phenotype of the parents.

Functional Annotation of All Salmonid Genomes (FAASG): An international initiative supporting future salmonid research, conservation and aquaculture

We describe an emerging initiative - the ‘Functional Annotation of All Salmonid Genomes’ (FAASG), which will leverage the extensive trait diversity that has evolved since a whole genome duplication event in the salmonid ancestor, to develop an integrative understanding of the functional genomic basis of phenotypic variation. The outcomes of FAASG will have diverse applications, ranging from improved understanding of genome evolution, to improving the efficiency and sustainability of aquaculture production, supporting the future of fundamental and applied research in an iconic fish lineage of major societal importance.

SalmonDB: a bioinformatics resource for Salmo salar and Oncorhynchus mykiss

SalmonDB is a new multiorganism database containing EST sequences from Salmo salar, Oncorhynchus mykiss and the whole genome sequence of Danio rerio, Gasterosteus aculeatus, Tetraodon nigroviridis, Oryzias latipes and Takifugu rubripes, built with core components from GMOD project, GOPArc system and the BioMart project. The information provided by this resource includes Gene Ontology terms, metabolic pathways, SNP prediction, CDS prediction, orthologs prediction, several precalculated BLAST searches and domains. It also provides a BLAST server for matching user-provided sequences to any of the databases and an advanced query tool (BioMart) that allows easy browsing of EST databases with user-defined criteria. These tools make SalmonDB database a valuable resource for researchers searching for transcripts and genomic information regarding S. salar and other salmonid species. The database is expected to grow in the near feature, particularly with the S. salar genome sequencing project. Database URL: http://genomicasalmones.dim.uchile.cl/