Correction to: L_RNA_scaffolder: scaffolding genomes with transcripts.

Following the publication of this article [1], the authors reported that the link to the software described in the article is no longer valid.

The transcriptomes of the crucian carp complex (Carassius auratus) provide insights into the distinction between unisexual triploids and sexual diploids.

Both sexual reproduction and unisexual reproduction are adaptive strategies for species survival and evolution. Unisexual animals have originated largely by hybridization, which tends to elevate their heterozygosity. However, the extent of genetic diversity resulting from hybridization and the genomic differences that determine the type of reproduction are poorly understood. In Carassius auratus, sexual diploids and unisexual triploids coexist. These two forms are similar morphologically but differ markedly in their modes of reproduction. Investigation of their genomic differences will be useful to study genome diversity and the development of reproductive mode. We generated transcriptomes for the unisexual and sexual populations. Genes were identified using homology searches and an ab initio method. Estimation of the synonymous substitution rate in the orthologous pairs indicated that the hybridization of gibel carp occurred 2.2 million years ago. Microsatellite genotyping in each individual from the gibel carp population indicated that most gibel carp genes were not tri-allelic. Molecular function and pathway comparisons suggested few gene expansions between them, except for the progesterone-mediated oocyte maturation pathway, which is enriched in gibel carp. Differential expression analysis identified highly expressed genes in gibel carp. The transcriptomes provide information on genetic diversity and genomic differences, which should assist future studies in functional genomics.

L_RNA_scaffolder: scaffolding genomes with transcripts.

BACKGROUND: Generation of large mate-pair libraries is necessary for de novo genome assembly but the procedure is complex and time-consuming. Furthermore, in some complex genomes, it is hard to increase the N50 length even with large mate-pair libraries, which leads to low transcript coverage. Thus, it is necessary to develop other simple scaffolding approaches, to at least solve the elongation of transcribed fragments. RESULTS: We describe L_RNA_scaffolder, a novel genome scaffolding method that uses long transcriptome reads to order, orient and combine genomic fragments into larger sequences. To demonstrate the accuracy of the method, the zebrafish genome was scaffolded. With expanded human transcriptome data, the N50 of human genome was doubled and L_RNA_scaffolder out-performed most scaffolding results by existing scaffolders which employ mate-pair libraries. In these two examples, the transcript coverage was almost complete, especially for long transcripts. We applied L_RNA_scaffolder to the highly polymorphic pearl oyster draft genome and the gene model length significantly increased. CONCLUSIONS: The simplicity and high-throughput of RNA-seq data makes this approach suitable for genome scaffolding. L_RNA_scaffolder is available at http://www.fishbrowser.org/software/L_RNA_scaffolder.

[Quantitative trait locus analysis of standard length, body depth and body thickness in mirror carp (Cyprinus carpio L.)].

Based on a full-sib family, the genetic linkage map was constructed with 246 microsatellite and 306 SNP markers, which was used to detect the QTLs for standard length (SL), body depth (H), body thickness (BT), and the ratio of standard length and body depth (SLH) in mirror carp by GridQTL software. The results indicated that a total of 14 related QTLs distributed on the 7 linkage groups were obtained. Seven QTLs were related to standard length, of which the linkage groups of LG6, LG17, LG21, LG23, and LG35 were at 5% significant level, and linkage group LG1 and LG28 were at 1% significant level, which explained 6.6%-12.6% of the phenotypic variance. Three QTLs were identified for body depth on the linkage groups of LG17, LG23 and LG28 (P amp; 0.01), accounting for 11.6%, 12.7%, and 15.6% of the phenotypic variance, respectively. Two QTLs were associated with body thickness on the linkage of LG23 and LG28 (P amp; 0.05), which explained 8.6% and 7.2% of the phenotypic variation, respectively. Two QTLs were responsible for the ratio of standard length and body depth on the linkage of LG21 and LG35 (P amp; 0.05), both of which explained 8.2% of the phenotypic variance. The results provide a useful reference for further candidate gene research and molecular marker assisted selection in mirror carp.

De novo assembly and characterization of the Hucho taimen transcriptome.

Taimen (Hucho taimen) is an important ecological and economic species that is classified as vulnerable by the IUCN Red List of Threatened Species; however, limited genomic information is available on this species. RNA-Seq is a useful tool for obtaining genetic information and developing genetic markers for nonmodel species in addition to its application in gene expression profiling. In this study, we performed a comprehensive RNA-Seq analysis of taimen. We obtained 157 M clean reads (14.7 Gb) and used them to de novo assemble a high-quality transcriptome with a N50 size of 1,060 bp. In the assembly, 82% of the transcripts were annotated using several databases, and 14,666 of the transcripts contained a full open reading frame. The assembly covered 75% of the transcripts of Atlantic salmon and 57.3% of the protein-coding genes of rainbow trout. To learn about the genome evolution, we performed a systematic comparative analysis across 11 teleosts including eight salmonids and found 313 unique gene families in taimen. Using Atlantic salmon and rainbow trout transcriptomes as the background, we identified 250 positive selection transcripts. The pathway enrichment analysis revealed a unique characteristic of taimen: It possesses more immune-related genes than Atlantic salmon and rainbow trout; moreover, some genes have undergone strong positive selection. We also developed a pipeline for identifying microsatellite marker genotypes in samples and successfully identified 24 polymorphic microsatellite markers for taimen. These data and tools are useful for studying conservation genetics, phylogenetics, evolution among salmonids, and selective breeding for threatened taimen.

JDP2: An oncogenic bZIP transcription factor in T cell acute lymphoblastic leukemia.

A substantial subset of patients with T cell acute lymphoblastic leukemia (T-ALL) develops resistance to steroids and succumbs to their disease. JDP2 encodes a bZIP protein that has been implicated as a T-ALL oncogene from insertional mutagenesis studies in mice, but its role in human T-ALL pathogenesis has remained obscure. Here we show that JDP2 is aberrantly expressed in a subset of T-ALL patients and is associated with poor survival. JDP2 is required for T-ALL cell survival, as its depletion by short hairpin RNA knockdown leads to apoptosis. Mechanistically, JDP2 regulates prosurvival signaling through direct transcriptional regulation of MCL1. Furthermore, JDP2 is one of few oncogenes capable of initiating T-ALL in transgenic zebrafish. Notably, thymocytes from rag2:jdp2 transgenic zebrafish express high levels of mcl1 and demonstrate resistance to steroids in vivo. These studies establish JDP2 as a novel oncogene in high-risk T-ALL and implicate overexpression of MCL1 as a mechanism of steroid resistance in JDP2-overexpressing cells.

The genetic map of goldfish (Carassius auratus) provided insights to the divergent genome evolutions in the Cyprinidae family.

A high-density linkage map of goldfish (Carassius auratus) was constructed using RNA-sequencing. This map consists of 50 linkage groups with 8,521 SNP markers and an average resolution of 0.62 cM. Approximately 84% of markers are in protein-coding genes orthologous to zebrafish proteins. We performed comparative genome analysis between zebrafish and medaka, common carp, grass carp, and goldfish to study the genome evolution events in the Cyprinidae family. The comparison revealed large synteny blocks among Cyprinidae fish and we hypothesized that the Cyprinidae ancestor undergone many inter-chromosome rearrangements after speciation from teleost ancestor. The study also showed that goldfish genome had one more round of whole genome duplication (WGD) than zebrafish. Our results illustrated that most goldfish markers were orthologous to genes in common carp, which had four rounds of WGD. Growth-related regions and genes were identified by QTL analysis and association study. Function annotations of the associated genes suggested that they might regulate development and growth in goldfish. This first genetic map enables us to study the goldfish genome evolution and provides an important resource for selective breeding of goldfish.

Complete mitochondrial genome sequence of Romanogobio tenuicorpus (Amur whitefin gudgeon).

Amur whitefin gudgeon (Romanogobio tenuicorpus) belongs to the family Cyprinidae, it is freshwater aquaculture species in China. In the report, we determined the complete mitochondrial genome sequence of Romanogobio tenuicorpus, which is 16,600 bp long circular molecule with 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes and a control region, the conserved sequence blocks, CSB1, CSB2 and CSB3 were also detected.

The complete mitochondrial genome sequence of Hemiculter leucisculus.

The complete mitochondrial genome of Hemiculter leucisculus was determined to be 16,617 bp. It contains the structure of 22 transfer RNA genes, 13 protein-coding genes, 2 ribosomal RNA genes, and non-coding control region (D-loop). The critical central conserved sequences (CSB-D, CSB-E, and CSB-F) were also detected. The determination of H. leucisculus mitogenome would play an important role in genetic diversity and population vitality in Cyprinidae.

The complete mitochondrial genome sequence of pike perch (Sander canadensis).

Pike perch (Sander canadensis) is a member of the largest order of Osteichthyes, Perciformes, and is an important ecological and economic freshwater species, which distributes in Ili River and Ergis River of Xinjiang Province, China. In this study, we sequenced the whole mitochondrial genome of pike perch, and analyzed the similarity with its related species. The mitochondrial genome of S. canadensis is 16,542 bp in length with 55.05% AT content, contained 13 protein coding genes, 22 tRNA genes, 2 ribosomal genes and an 892 bp non-coding region. In control region, 6 CSBs (CSB-1, CSB-2, CSB-3, CSB-D, CSB-E and CSB-F), one potential TAS and one poly-T region were identified. Comparing all protein-coding genes and whole genome sequence with 4 species of Perciformes (three species of Percidae, Perca flavescens. Percina macrolepida. Etheostoma radiosum and one outgroup Oreochromis sp. red tilapia), ND3 gene has the highest mutation rate, and S. canadensis has higher similarity with Perca flavescens than others. The mitochondrial genomic sequence will help us to study the conservation genetic and evolution of Percidae.

A zebrafish model of myelodysplastic syndrome produced through tet2 genomic editing.

The ten-eleven translocation 2 gene (TET2) encodes a member of the TET family of DNA methylcytosine oxidases that converts 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) to initiate the demethylation of DNA within genomic CpG islands. Somatic loss-of-function mutations of TET2 are frequently observed in human myelodysplastic syndrome (MDS), which is a clonal malignancy characterized by dysplastic changes of developing blood cell progenitors, leading to ineffective hematopoiesis. We used genome-editing technology to disrupt the zebrafish Tet2 catalytic domain. tet2(m/m) (homozygous for the mutation) zebrafish exhibited normal embryonic and larval hematopoiesis but developed progressive clonal myelodysplasia as they aged, culminating in myelodysplastic syndromes (MDS) at 24 months of age, with dysplasia of myeloid progenitor cells and anemia with abnormal circulating erythrocytes. The resultant tet2(m/m) mutant zebrafish lines show decreased levels of 5hmC in hematopoietic cells of the kidney marrow but not in other cell types, most likely reflecting the ability of other Tet family members to provide this enzymatic activity in nonhematopoietic tissues but not in hematopoietic cells. tet2(m/m) zebrafish are viable and fertile, providing an ideal model to dissect altered pathways in hematopoietic cells and, for small-molecule screens in embryos, to identify compounds with specific activity against tet2 mutant cells.

The Complete mitochondrial genome of Spinibarbus denticulatus (Oshima).

Spinibarbus denticulatus (Oshima) is a rare and commercial fish. The complete mitochondrial genome sequence of S. denticulatus will help us to study the genetic of conversation, such as the genetic diversity and genetic structure, and provides the basis for the study in evolution. In this paper, the complete mitochondrial genome of S. denticulatus was determined to be 16,549 bp in length by Sanger sequencing technology. Thirteen protein-coding genes, 22 tRNA genes and 2 ribosomal genes were characterized. We also analyzed the structure of control region, 6 CSBs (CSB-1, CSB-2, CSB-3, CSB-D, CSB-E and CSB-F) and 1 TAS were identified, the control region also included an AT unit tandem repeat with 17 repeat times.

Mitochondrial DNA sequence of Mongolian redfin (Chanodichthys mongolicus).

Monglian redfin is a kind of freshwater aquaculture species which has an important economic value in China. In this study, we report the complete sequence of mitochondrial genome of Monglian redfin. The complete mitochondrial genome sequence is determined to be 16,621 base pairs (bp) in length and contain 13 protein-coding gene (PCGs), 22 transfer RNA genes, large (rrnL) and small (rrnS) ribosomal RNA, and the non-coding control region. Its total A+T content is 55.98%. We also analyzed the structure of control region, six conserved sequence blocks (CSBs) (CSB-1, CSB-2, CSB-3, CSB-D, CSB-E and CSB-F) and one potential termination-associated sequence were detected and the control region also included a 2-bp tandem repeat with eight repeat times.

Duplication and differentiation of common carp (Cyprinus carpio) myoglobin genes revealed by BAC analysis.

Two distinct myoglobin (mb) transcripts have been reported in common carp, Cyprinus carpio, which is a hypoxia-tolerant fish living in habitats with greatly fluctuant dissolved oxygen levels. Recombinant protein analysis has shown functional specialization of the two mb transcripts. In this work, analysis for mb-containing bacterial artificial chromosome (BAC) clones indicated different genome loci for common carp myoglobin-1 (mb-1) and myoglobin-2 (mb-2) genes. Fluorescence in situ hybridization (FISH) revealed that mb-1 and mb-2 are located on separate chromosomes. In both of the mb-1 and mb-2 containing BAC clones, gene synteny was well conserved with the homologous region on zebrafish chromosome 1, supporting that the common carp specific mb-2 gene originated from the recent whole genome duplication event in cyprinid lineage. Transcription factor binding sites search indicated that both common carp mb genes lacked specificity Protein 1 (Sp1) and myocyte enhancer factor-2 (MEF2) binding sites, which mediated muscle-specific and calcium-dependent expression in the well-studied mb promoters. Potential hypoxia response elements (HREs) were predicted in the regulatory region of common carp mb genes. These characteristics of common carp mb gene regulatory region well interpreted the hypoxia-inducible, non-muscle expression pattern of mb-1. In the case of mb-2, a 10 bp insertion to the binding site of upstream stimulatory factor (USF), which was a co-factor of hypoxia inducible factor (HIF), might cause the non-response to hypoxia treatment of mb-2. The case of common carp mb gene duplication and subsequent differentiation in expression pattern and protein function provided an example for adaptive evolution toward aquatic hypoxia tolerance.

[Screening and body correlation analysis of microsatellite markers related to intermuscular bone number in common carp (Cyprinus carpio)].

In this study, 149 polymorphic markers were screened from 200 microsatellite markers. From a family of mirror carp, which included 107 individuals. All samples were analyzed for body correlation, and intermuscular bone number was tested using the General Linear Model (GLM) single marker regression. Determination of the threshold values by 10,000 permutation tests showed that eight markers had significant correlation (P<0.05), in which HLJ3086, HLJ3642 and HLJ3515 had very significant correlation with intermuscular bone number (P<0.01). In addition, the genotypes of the captured correlative loci were determined by Duncan's test using SPSS17.0 software. Markers were used to screen the protein and nucleotide database in the National Center for Biotechnology Information (NCBI). Blasting results showed that HLJ2891 was highly correlated (92%) with latrophilin-2-like and HLJ3515 was highly correlated (81%) with serine/threonine-protein kinase 32B-like of zebrafish. These functional markers and genotypes may provide an efficient basis for marker-assisted selection of intermuscular bone number in mirror carp.

[Analysis of genetic diversity on 9 wild stocks of taimen (Hucho taimen) by microsatellite markers].

Taimen (Hucho taimen) is a native fish species in China and it is in the state of endangerment. To explain clearly the genetic diversity and genetic structure, 9 wild populations of taimen were investigated using 20 microsatellite markers. The results showed that their observed heterozygosity ranged from 0.0994 to 0.8882, the expected heterozygosity varied from 0.2005 to 0.8759, and the range of PIC index was from 0.3432 to 0.5261 while population from Huma River had low genetic diversity. Fst of matching group ranged from 0.0246 to 0.2333 (P <0.0001)and Nm varied among 0.8216 to 9.9292, which indicated that the genetic differentiation was remarkable among populations.The half/full-sib family tests detected a proportion of half/full-sib family groups varying among 27.78% to 90.91%, showing a high inbred pressure and a risk of bottlenecks experienced by most groups. The AMOVA results showed that the global Fst was 0.1081; the clustering result showed that individuals from Beiji tributary of Heilongjiang River clustered as one clade, all individuals from Huma River and Wusuli River clustered as one clade and all individuals from the upper reaches of the Heilongjiang River clustered as another clade. All these results indicated that the decrease of taimen resource has affected the gene exchange among their populations. In order to achieve full protection of taimen germplasm resources, we should put an end to the destructive fishing for taimen and promotegene exchange among their populations.

[Applicability of microsatellite DNA markers to the parental identification of Hucho taimen (Pallas)].

Using seven pairs of microsatellite markers we studied the relationship between the size of the the candidate groups and the microsatellite paternity testing accuracy by separate breed or the microsatellite paternity appraisal ability by mixed breed. By computer software analysis, the appraisal ability decreased with the candidate groups increase. For the 81 putative parents the microsatellite paternity testing accuracy was 80% and the microsatellite paternity appraisal ability was 78.9%; for the 9 putative parents, the testing accuracy was 93.3% and the appraisal ability was 92.2%. The results indicated that the microsatellite DNA markers can be used for the parentage determination of Hucho taimen Pallas.