BackgroundHigh density linkage maps are essential for comparative analysis of synteny, fine mapping of quantitative trait loci (QTL), searching for candidate genes and facilitating genome sequence assembly. However, in most foodfish species, marker density is still low. We previously reported a first generation linkage map with 240 DNA markers and its application to preliminarily map QTL for growth traits in Asian seabass (Lates calcarifer). Here, we report a high-resolution linkage map with 790 microsatellites and SNPs, comparative analysis of synteny, fine-mapping of QTL and the identification of potential candidate genes for growth traits.ResultsA second generation linkage map of Asian seabass was developed with 790 microsatellite and SNP markers. The map spanned a genetic length of 2411.5 cM, with an average intermarker distance of 3.4 cM or 1.1 Mb. This high density map allowed for comparison of the map with Tetraodon nigroviridis genome, which revealed 16 synteny regions between the two species. Moreover, by employing this map we refined QTL to regions of 1.4 and 0.2 cM (or 400 and 50 kb) in linkage groups 2 and 3 in a population containing 380 progeny; potential candidate genes for growth traits in QTL regions were further identified using comparative genome analysis, whose effects on growth traits were investigated. Interestingly, a QTL cluster at Lca371 underlying growth traits of Asian seabass showed similarity to the cathepsin D gene of human, which is related to cancer and Alzheimer's disease.ConclusionsWe constructed a high resolution linkage map, carried out comparative mapping, refined the positions of QTL, identified candidate genes for growth traits and analyzed their effects on growth. Our study developed a framework that will be indispensable for further identification of genes and analysis of molecular variation within the refined QTL to enhance understanding of the molecular basis of growth and speed up genetic improvement of growth performance, and it also provides critical resource for future genome sequence assembly and comparative genomics studies on the evolution of fish genomes.

Barramundi (Lates calcarifer) is an important farmed marine food fish species. Its compact genome ($700 Mb) is among the smallest genomes of food fish species. We established a first-generation genetic linkage map of Barramundi with a mapping panel containing three parents (two males and one female) and 93 progeny. A total of 240 microsatellite markers were mapped into 24 linkage groups. Among these markers, 10 were located in ESTs and known genes. The total lengths of the female and male maps were 873.8 and 414.5 cM with an average marker spacing of 6.20 and 4.70 cM, respectively. Comparing the flanking sequences of the 240 Barramundi microsatellites with the assembled whole-genome sequences of Tetraodon nigrovidiris revealed 55 homologous sequences located in 19 of the 21 chromosomes of T. nigrovidiris. The map will not only enable the mapping of quantitative trait loci, but also provide new resources for understanding the evolution of fish genomes.

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High genetic diversity is thought to characterize successful invasive species, as the potential to adapt to new environments is enhanced and inbreeding is reduced. The red swamp crayfish, Procambarus clarkii, native to northeastern Mexico and southcentral USA was introduced to Nanjing, China from Japan in 1929. Little is known about the genetic diversity and population structure of this species in China. We examined the genetic diversity and population structure of six P. clarkii populations using nine polymorphic microsatellites. Among the six populations, Nanjing population showed the highest allele number, allele richness and gene diversity, which is consistent with records indicating Nanjing may be the first site of introduction. In all six populations, significant heterozygote deficit was observed, suggesting founder effects and non-random mating. Analysis of bottleneck under infinite allele model, stepwise mutation model and two-phased model of mutation revealed evidence of a recent bottleneck in all these populations. Pairwise genetic distance analysis, AMOVA and assignment tests demonstrated high genetic differentiation between populations. Pairwise genetic distance did not fit the pairwise geographic distance, suggesting that human mediated dispersal have played a role in the population expansion and genetic differentiation.

Jatropha curcas is a potential plant species for biodiesel production. However, its seed yield is too low for profitable production of biodiesel. To improve the productivity, genetic improvement through breeding is essential. A linkage map is an important component in molecular breeding. We established a first-generation linkage map using a mapping panel containing two backcross populations with 93 progeny. We mapped 506 markers (216 microsatellites and 290 SNPs from ESTs) onto 11 linkage groups. The total length of the map was 1440.9 cM with an average marker space of 2.8 cM. Blasting of 222 Jatropha ESTs containing polymorphic SSR or SNP markers against EST-databases revealed that 91.0%, 86.5% and 79.2% of Jatropha ESTs were homologous to counterparts in castor bean, poplar and Arabidopsis respectively. Mapping 192 orthologous markers to the assembled whole genome sequence of Arabidopsis thaliana identified 38 syntenic blocks and revealed that small linkage blocks were well conserved, but often shuffled. The first generation linkage map and the data of comparative mapping could lay a solid foundation for QTL mapping of agronomic traits, marker-assisted breeding and cloning genes responsible for phenotypic variation.

Asian sea bass (Lates calcarifer) is one of the most important marine food ¢sh species in Asia and Australia. To estimate the reproductive success of broodstock and heritabilities of growth-related traits, two independent full-factorial crosses (PI and PII) were created by crossing 10 males and 10 females. At 90 days post hatch (dph), the body weight (BW) and total length (TL) of 804 individuals from PI and 900 individuals from PII were measured and tissues samples of each ¢sh were collected. Parents and o¡spring were genotyped with nine polymorphic microsatellites. Out of 1704 o¡spring from the two crosses, 98.7% were assigned to their parents. In PI, 19 of 20 parents produced o¡spring, while in PII, only ¢ve parents contributed to o¡spring. Low contribution of parents to o¡spring could lead to reduced genetic variation in the next generation. Heritabilities for growth-related traits were estimated using the pedigrees reconstructed using microsatellite genotypes. The estimates of heritability were 0.22 AE 0.16 and 0.25 AE 0.18 for BW, 0.31 AE 0.14 and 0.24 AE 0.21 for TL and 0.22 AE 0.22 and 0.15 AE 0.09 for Fulton's condition factor in the two crosses respectively. Body weights at 90 dph and at harvest (289 dph) were sig-ni¢cantly correlated (r 5 0.601, Po0.01). Therefore, growth-related traits could be improved by exploiting additive genetic e¡ects through selective breeding, and broodstock candidates could be selected early in the production cycle. BW, body weight; TL, total length; K, Fulton's condition factor. ÃÃ Signi¢cance at P 5 0.01.Reproduction and heritability of growth traits in Lates calcarifer C M Wang et al.

BackgroundThe Asian seabass (Lates calcarifer) is an important marine foodfish species in Southeast Asia and Australia. Genetic improvement of this species has been achieved to some extent through selective breeding programs since 1990s. Several genomic tools such as DNA markers, a linkage map, cDNA and BAC libraries have been developed to assist selective breeding. A physical map is still lacking, although it is essential for positional cloning of genes located in quantitative trait loci (QTL) and assembly of whole genome sequences.Methodology/Principal FindingsA genome-wide physical map of the Asian seabass was constructed by restriction fingerprinting of 38,208 BAC clones with SNaPshot HICF FPC technique. A total of 30,454 were assembled into 2,865 contigs. The physical length of the assembled contigs summed up to 665 Mb. Analyses of some contigs using different methods demonstrated the reliability of the assembly.Conclusions/SignificanceThe present physical map is the first physical map for Asian seabass. This physical map will facilitate the fine mapping of QTL for economically important traits and the positional cloning of genes located in QTL. It will also be useful for the whole genome sequencing and assembly. Detailed information about BAC-contigs and BAC clones are available upon request.

Parvalbumins are extremely abundant in fish muscle and play an important role in muscle relaxation. In this study, two parvalbumin genes (PVALB1 and PVALB2) were cloned from Asian seabass (Lates calcarifer). The cDNAs for PVALB1 and PVALB2 were 840 and 667 bp respectively. Both genes consisted of five exons and four introns, encoded 109 amino acids, and were of beta lineage. Using real-time polymerase chain reaction, expression of PVALB1 was detected in all 10 tissues tested, with expression in brain, kidney, muscle and small intestine being 15- to 322-fold higher than in the other tissues. Expression of PVALB2 was detected only in muscle, brain and intestine, and was up to 10-fold lower than PVALB1 expression. A (CT)(17) microsatellite in the 3'-untranslated region of PVALB1 and three single nucleotide polymorphisms (SNPs) in the third intron of PVALB2 were identified. The microsatellite in PVALB1 was significantly associated with body weight and body length at 90 days post-hatch (P < 0.01), whereas the SNPs in PVALVB2 were not associated with these traits.