Effects of Gene Alternative Splicing Events on Resistance to Cryptocaryonosis of Large Yellow Croaker (Larimichthys crocea).

Large yellow croaker (L. crocea) is a productive species in marine aquaculture with great economic value in China. However, the sustainable development of large yellow croaker is hampered by various diseases including cryptocaryonosis caused by Cryptocaryon irritans. The genetic regulation processes for cryptocaryonosis in large yellow croaker are still unclear. In this present study, we analyzed differential alternative splicing events between a C. irritans resistance strain (RS) and a commercial strain (CS). We identified 678 differential alternative splicing (DAS) events from 453 genes in RS and 719 DAS events from 500 genes in CS. A set of genes that are specifically alternatively spliced in RS was identified including mfap5, emp1, and trim33. Further pathway analysis revealed that the specifically alternative spliced genes in RS were involved in innate immune responses through the PRR pathway and the Toll and Imd pathway, suggesting their important roles in the genetic regulation processes for cryptocaryonosis in large yellow croaker. This study would be helpful for the studies of the pathogenesis of cryptocaryonosis and dissection of C. irritans resistance for L. crocea.

Genomic Selection of Large Yellow Croaker (Larimichthys crocea) with a High Plant Protein Diet Enhances the Growth Performance of Offspring.

Fishmeal is over-represented in the diets of large yellow croaker (Larimichthys crocea), and this farming mode, which relies heavily on fishmeal, is highly susceptible to the price of fishmeal and is unsustainable. Therefore, more and more studies on the large yellow croaker tend to replace fishmeal with land-based animal or plant proteins, but few studies have considered it from the genomic selection. In this study, we evaluated the survival rate (SR), final body weight (FBW), body weight gain (BWG), weight gain rate (WGR), and specific growth rate (SGR) of the large yellow croaker GS7 strain, which was obtained through genomic selection for tolerance to plant proteins and analyzed the differences in plant protein utilization between the GS7 strain and unselected commercial large yellow croaker (control group). The results of separate feeding for 60 days showed that although there was no significant difference in SR between the control and GS7 strains (P > 0.05), the BWG, WGR, and SGR of the control were significantly lower (P < 0.05) than those of the GS7 group. Results of mixed feeding after PIT marking showed that compared to the control fish, the GS7 strain had significantly higher BWG, WGR, and SGR (P < 0.0001). To make the experimental results more precise, we compared fishes with equivalent initial body weight (IBW) in the GS7 strain and the control group. The final fish body weight (FBW) of Ctrl-2 (IBW 300-399 g) and Ctrl-4 (IBW 500-599 g) was significantly lower than those of the corresponding GS7-2 and GS7-4 (P < 0.05), while the FBW of Ctrl-1 (IBW 200-299 g) and Ctrl-3 (IBW 400-499 g) was much significantly lower than the corresponding GS7-1 and GS7-3 (P < 0.01). The BWG, WGR, and SGR of Ctrl-1 and Ctrl-4 were more significantly lower than those of the corresponding GS7-1 and GS7-4 (P < 0.01), while the BWG, WGR, and SGR of Ctrl-2 and Ctrl-3 were more significantly different from the corresponding GS7-2 and GS7-3 (P < 0.0001). Our results seem to point toward the same conclusion that the GS7 strain is better adapted to high plant protein diets than the unselected commercial large yellow croaker. These results will provide a reference for the low-fishmeal culture industry of large yellow croakers and the selection and breeding of strains tolerant to a high percentage of plant proteins in other marine fishes.

Non-invasive methods for heart rate measurement in fish based on photoplethysmography.

Heart rate is a crucial physiological indicator for fish, but current measurement methods are often invasive or require delicate manipulation. In this study, we introduced two non-invasive and easy-to-operate methods based on photoplethysmography, namely reflectance-type photoplethysmography (PPG) and remote photoplethysmography (rPPG), which we applied to the large yellow croaker (Larimichthys crocea). PPG showed perfect synchronization with electrocardiogram (ECG), with a Pearson's correlation coefficient of 0.99999. For rPPG, the results showed good agreement with ECG. Under active provision of green light, the Pearson's correlation coefficient was 0.966, surpassing the value of 0.947 under natural light. Additionally, the root mean square error was 0.810, which was lower than the value of 1.30 under natural light, indicating not only that the rPPG method had relatively high accuracy but also that green light may have the potential to further improve its accuracy.

First Genomic Prediction of Single-Step Models in Large Yellow Croaker.

Genome selection is mainly used in disease-resistant traits of aquatic species; however, its implementation is hindered by a high cost of genotype and phenotype data collection. Single-step genomic best linear unbiased prediction (SSGBLUP) can integrate phenotypes, genetic markers, and pedigree records into simultaneous prediction without increasing genotyping costs. The objective of this study is to investigate the performance of SSGBLUP in large yellow croaker and to evaluate the effects of the number of phenotypic records and genotyping per family on the predictive ability of SSGBLUP. A large yellow croaker population consists of 6898 individuals from 14 families with survival time resistant against Cryptocaryon irritans (C. irritans), body weight (BW), and body length (BL) traits were collected, of which 669 individuals were genotyped. Results showed that the mean predictive ability of all traits in the individuals randomly sampling for SSGBLUP, GBLUP, and BLUP was 0.738, 0.738, and 0.736, respectively. Moreover, the predictive ability of SSGBLUP and BLUP models did not increase with the extra phenotypic records per family, in which the predictive ability of SSGBLUP and BLUP in survival time was 0.853 and 0.851 for only genotyped data (N = 0) used, and 0.852, 0.845 for all phenotypic records (N = 600) used, respectively. However, with the increase in the genotype number of training set, the prediction ability of SSGBLUP and GBLUP model was increased and the highest predictive ability was gained when the genotype number per family was 40 or 45. In addition, the prediction ability of SSGBLUP model was higher than that of GBLUP. Our study showed that the SSGBLUP model still has great potential and advantages in genomic breeding of large yellow croakers. It is recommended that each family provide 100 phenotypic individuals, of which 40 individuals with genotyping data for SSGBLUP model prediction and family resistance evaluation.

Population structure and genome-wide evolutionary signatures reveal putative climate-driven habitat change and local adaptation in the large yellow croaker.

The large yellow croaker (Larimichthys crocea) is one of the most economically valuable marine fish in China and is a notable species in ecological studies owing to a serious collapse of wild germplasm in the past few decades. The stock division and species distribution, which have important implications for ecological protection, germplasm recovery, and fishery resource management, have been debated since the 1960s. However, it is still uncertain even how many stocks exist in this species. To address this, we evaluated the fine-scale genetic structure of large yellow croaker populations distributed along the eastern and southern Chinese coastline based on 7.64 million SNP markers. Compared with the widely accepted stock boundaries proposed in the 1960s, our results revealed that a climate-driven habitat change probably occurred between the Naozhou (Nanhai) Stock and the Ming-Yuedong (Mindong) Stock. The boundary between these two stocks might have shifted northwards from the Pearl River Estuary to the northern area of the Taiwan Strait, accompanied by highly asymmetric introgression. In addition, we found divergent landscapes of natural selection between the stocks inhabiting northern and southern areas. The northern population exhibited highly agminated signatures of strong natural selection in genes related to developmental processes, whereas moderate and interspersed selective signatures were detected in many immune-related genes in the southern populations. These findings establish the stock status and genome-wide evolutionary landscapes of large yellow croaker, providing a basis for conservation, fisheries management and further evolutionary biology studies. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-023-00165-2.

Integrated mRNA and miRNA expression analyses for Cryptocaryon irritans resistance in large yellow croaker (Larimichthys crocea).

Large yellow croaker (Larimichthys crocea) is one of the most important mariculture fish in China. However, cryptocaryonosis caused by Cryptocryon irritans infection has brought huge economic losses and threatened the healthy and sustainable development of L. crocea industry. Recently, a new C. irritans resistance strain of L. crocea (RS) has been bred using genomic selection technology in our laboratory work. However, the molecular mechanisms for C. irritans resistance of RS have not been fully understood. MicroRNAs (miRNAs) are endogenous small non-coding RNAs that are post-transcriptional regulators, and they play vital roles in immune process of bony fish. Identification of anti-C.irritans relevant miRNA signatures could, therefore, be of tremendous translational value. In the present study, integrated mRNA and miRNA expression analysis was used to explore C. irritans resistance mechanisms of the L. crocea. RS as well as a control strain (CS) of L. crocea, were artificially infected with C. irritans for 100 h, and their gill was collected at 0 h (pre-infection), 24 h (initial infection), and 72 h (peak infection) time points. The total RNA from gill tissues was extracted and used for transcriptome sequencing and small RNA sequencing. After sequencing, 23,172 known mRNAs and 289 known miRNAs were identified. The differential expression was analyzed in these mRNAs and mRNAs and the interactions of miRNA-mRNA pairs were constructed. KEGG pathway enrichment analyses showed that these putative target mRNAs of differentially expressed miRNAs (DEMs) were enriched in different immune-related pathways after C. irritans infection in RS and CS. Among them, necroptosis was the immune-related pathway that was only significantly enriched at two infection stages of RS group (RS-24 h/RS-0h and RS-72 h/RS-0h). Further investigation indicates that necroptosis may be activated by DEMs such as miR-133a-3p, miR-142a-3p and miR-135c, this promotes inflammation responses and pathogen elimination. These DEMs were selected as miRNAs that could potentially regulate the C. irritans resistance of L. crocea. Though these inferences need to be further verified, these findings will be helpful for the research of the molecular mechanism of C. irritans resistance of L. crocea and miRNA-assisted molecular breeding of aquatic animals.

Identification and Expression Analysis of LncRNAs Reveal the Immune Mechanism of Visceral White-Nodules Disease Resistance in Large Yellow Croaker.

Long non-coding RNAs (lncRNAs) have several known functions in fish growth processes and signal transduction, but their possible roles in response to bacterial diseases remain largely unresolved. In this study, we report a comprehensive cold-water bacterial disease-responsive lncRNA expression profile for understanding the transcriptional regulatory mechanisms of visceral white-nodules disease resistance in large yellow croaker. A total of 2534 high-confidence lncRNAs were identified by a rigorous filtering pipeline as a basic sequence set for comparative transcriptional analysis. In addition, a total of 10,200 lncRNA-mRNA pairs with high correlation coefficients were identified by expressions level correlation analysis, including non-redundant 381 DE lncRNAs and 2590 differential expressed genes. MSTRG_11084_1 and MSTRG_20402_1 were linked to a large number of target genes and may be involved in important functions in immune regulation. We further revealed the conserved and idiosyncratic features of the disease response process between the technical control strain (TCS) and the resistant strain (RS). Immune-related pathways were enriched in GO terms and KEGG pathways, among which cytokine-cytokine receptor interaction, MAPK signaling pathway, and NF-kappa B signaling pathway may play a key role in VWND resistance in large yellow croaker. Protein-protein interaction network (PPI) analysis revealed that immune-related target genes such as il-10, met, acta2, myc, cav1, and ntrk1, as well as growth and metabolism-related target genes such as pik3r2, igf1, sc5d, hmgcr, and lss were considered the main hub genes. This study represents the first characterization of lncRNAs involved in VWND resistance in large yellow croaker and provides new clues for elucidating the disease response mechanism of large yellow croaker.

Integrative analysis of GWAS and transcriptome reveals p53 signaling pathway mediates resistance to visceral white-nodules disease in large yellow croaker.

Visceral white-nodules disease (VWND), caused by Pseudomonas plecoglossicida, is one of the primary causes of morbidity and mortality in large yellow croaker aquaculture. Host disease resistance is a heritable trait that involves complex regulatory processes. However, the regulatory mechanism of bacterial resistance in large yellow croaker is still unclear. This study attempted to systematically evaluate the major genetic loci and transcriptional regulatory mechanisms associated with the resistance to VWND in large yellow croaker by crossover method studies. A large population of large yellow croaker was challenged with P. plecoglossicida, with survival time recorded and samples were taken for genotyping. Meanwhile, spleen samples that were used for RNA-seq to compare their transcriptomic profiles before and after infection were taken from resistant populations (RS) and susceptible control populations (CS) bred using the genomic selection (GS) technique. Genome-wide association analyses using 46 K imputed SNP genotypes highlighted that resistance is a polygenic trait. The integrative analysis results show the co-localization of the cd82a gene between disease resistance-related genetic loci and comparative transcriptional analysis. And functional enrichment analysis showed differential enrichment of the p53 signaling pathway in RS and CS groups, suggesting that there may be cd82a-mediated p53 signaling pathway activation for VWND resistance. This large-scale study provides further evidence for the heritability and transcriptional regulatory mechanisms of host inheritance of VWND resistance.

Genome-Wide Association and Expression Analysis Revealed the Candidate Variants and Molecular Underpinnings of Cold-Stress Response in Large Yellow Croaker.

Large yellow croaker (Larimichthys crocea) is one of the most economically important fish in China. Recently, global climate change has caused more and more intense and extreme low temperature weathers, resulting in huge losses to the large yellow croaker industry. Therefore, it is essential to understand the mechanisms of low-temperature tolerance in large yellow croaker. Here, we conducted an integrative analysis of genome-wide association study (GWAS) and transcriptome analysis to identify candidate variants and reveal the molecular underpinning of cold-stress response in large yellow croaker. A total of 8 significant single nucleotide polymorphisms (SNPs) loci on 6 chromosomes were identified in the GWAS analysis, and 5764 (gill) and 3588 (liver) differentially expressed genes (DEGs) were detected in cold-stressed large yellow croaker, respectively. Further comparative and functional analysis of the candidate genes and DEGs highlighted the importance of pathways/genes related to immune response, cellular stress response, lipid transport, and metabolism in the cold-stress response of large yellow croaker. Our results provide insights into the cold tolerance of large yellow croaker and contribute to genomic-based selection for low-temperature-resistant large yellow croaker.

Disruption of mstn Gene by CRISPR/Cas9 in Large Yellow Croaker (Larimichthys crocea).

The large yellow croaker (Larimichthys crocea) plays an economically vital role in the marine aquaculture in China. Suffering from infection of bacteria and protozoon, effect of extreme weather and stress from high-density farming, genome editing is thought to be an important tool applied to L. croea for enhancing commercial traits such as growth rate, disease resistance, and nutrition component. In this study, we identified two mstn genes in L. croea and investigated the different phylogenetic clades, gene structures, and conserved syntenic relationships. To obtain fast-growing large yellow croaker, we specially selected two validated targets for mstnb knockout, which was homologous to mammalian myostatin gene (MSTN) and downregulated skeletal muscle growth and development. Five significant mutation types were generated in two mosaic mutants by transferring specific CRISPR/Cas9 RNPs (ribonucleoprotein) into the one-cell fertilized embryos based on CRISPR/Cas9 technology. Subsequently, we also elucidated the obstacles and possible measures to improve the success rate of inducing modified large yellow croaker. Our results would provide valuable method and reference for facilitating genome editing programs of the large yellow croaker in the future.

First genomic prediction and genome-wide association for complex growth-related traits in Rock Bream (Oplegnathus fasciatus).

Rock Bream (Oplegnathus fasciatus) is an important aquaculture species for offshore cage aquaculture and fish stocking of marine ranching in East Asia. Genomic selection has the potential to expedite genetic gain for the key target traits of a breeding program, but has not yet been evaluated in Oplegnathus. The purposes of the present study were to explore the performance of genomic selection to improve breeding value accuracy through real data analyses using six statistical models and to carry out genome-wide association studies (GWAS) to dissect the genetic architecture of economically vital growth-related traits (body weight, total length, and body depth) in the O. fasciatus population. After quality control, genotypes for 16,162 SNPs were acquired for 455 fish. Heritability was estimated to be moderate for the three traits (0.38 for BW, 0.33 for TL, and 0.24 for BD), and results of GWAS indicated that the underlying genetic architecture was polygenic. Six statistic models (GBLUP, BayesA, BayesB, BayesC, Bayesian Ridge-Regression, and Bayesian LASSO) showed similar performance for the predictability of genomic estimated breeding value (GEBV). The low SNP density (around 1 K selected SNP based on GWAS) is sufficient for accurate prediction on the breeding value for the three growth-related traits in the current studied population, which will provide a good compromise between genotyping costs and predictability in such standard breeding populations advanced. These consequences illustrate that the employment of genomic selection in O. fasciatus breeding could provide advantages for the selection of breeding candidates to facilitate complex economic growth traits.

Comparative transcriptome analysis reveals immunoregulation mechanism of lncRNA-mRNA in gill and skin of large yellow croaker (Larimichthys crocea) in response to Cryptocaryon irritans infection.

BACKGROUND: Cryptocaryonosis caused by Cryptocaryon irritans is one of the major diseases of large yellow croaker (Larimichthys crocea), which lead to massive economic losses annually to the aquaculture industry of L. crocea. Although there have been some studies on the pathogenesis for cryptocaryonosis, little is known about the innate defense mechanism of different immune organs of large yellow croaker. RESULTS: In order to analyze the roles of long non-coding RNAs and genes specifically expressed between immune organs during the infection of C. irritans, in this study, by comparing transcriptome data from different tissues of L. crocea, we identified tissue-specific transcripts in the gills and skin, including 507 DE lncRNAs and 1592 DEGs identified in the gills, and 110 DE lncRNAs and 1160 DEGs identified in the skin. Furthermore, we constructed transcriptome co-expression profiles of L. crocea gill and skin, including 7,503 long noncoding RNAs (lncRNAs) and 23,172 protein-coding genes. Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses showed that the DEGs and the target genes of the DE lncRNAs in the gill were specifically enriched in several pathways related to immune such as HIF-1 signaling pathway. The target genes of DE lncRNAs and DEGs in the skin are specifically enriched in the complement and coagulation cascade pathways. Protein-protein interaction (PPI) network analysis identified 3 hub genes including NFKBIA, TNFAIP3 and CEBPB, and 5 important DE lncRNAs including MSTRG.24134.4, MSTRG.3038.5, MSTRG.27019.3, MSTRG.26559.1, and MSTRG.10983.1. The expression patterns of 6 randomly selected differentially expressed immune-related genes were validated using the quantitative real-time PCR method. CONCLUSIONS: In short, our study is helpful to explore the potential interplay between lncRNAs and protein coding genes in different tissues of L. crocea post C. irritans and the molecular mechanism of pathogenesis for cryptocaryonosis. HIGHLIGHTS: Skin and gills are important sources of pro-inflammatory molecules, and their gene expression patterns are tissue-specific after C. irritans infection. 15 DEGs and 5 DE lncRNAs were identified as hub regulatory elements after C. irritans infection The HIF-1 signaling pathway and the complement and coagulation cascade pathway may be key tissue-specific regulatory pathways in gills and skin, respectively.

Sex-Specific Genomic Region Identification and Molecular Sex Marker Development of Rock Bream (Oplegnathus fasciatus).

Rock bream (Oplegnathus fasciatus) is a valuable commercial marine teleost species, which exhibits sexual dimorphism in growth performance. However, the absence of a rapid and cost-effective sex identification method based on sex-specific genetic marker has impeded study on sex determination mechanisms and breeding applications. In the present study, we firstly developed the PCR method for identifying potential sex-specific sequences in Oplegnathus fasciatus with the next-generation sequencing. Sex-specific genomic regions/loci for sex determination were discovered on Chr2 and Chr6 by genome-wide association analysis, sequencing depth, and heterozygosity comparison between females and males. Candidate sex-determining genes (CCDC63, ITR, WNT4) were furtherly detected in transcriptome data of testes and ovaries. Taken together, a male-specific 34-bp deletion on the Chr2 was identified and developed into molecular marker of sex for O. fasciatus. After validation in individuals with known phenotypic sexes, the accuracy was 100%. This study gives an insight into the mechanism of sex determination in O. fasciatus, and the gender marker is crucial both for future genomic research and for development of efficient and sustainable aquaculture practice.

Genome-Wide Association Analysis Reveals the Genetic Architecture of Parasite (Cryptocaryon irritans) Resistance in Large Yellow Croaker (Larimichthys crocea).

Large yellow croaker is an important marine culture species in China. Recently, the large yellow croaker industry is threatened by various disease problems, especially for the white spot disease, which is caused by parasite Cryptocaryon irritans. In the current study, we conducted a genome-wide association study (GWAS) for C. irritans resistance in two large yellow croaker populations (n = 264 and n = 480, respectively). We identified 15 QTL with explained genetic variance ranging from 1 to 8% in the two populations. One QTL on chromosome 23 was shared by the two populations, and three QTL had been reported in the previous study. We identified a lot of biological pathways associated with C. irritans resistance, such as hormone transport, response to bacterium, apoptotic process, acute inflammatory response to antigenic stimulus, and NF-kappa B signaling pathway. The genes casp8 and traf6 involved in regulatory network for apoptosis and inflammation were identified to be candidate genes for C. irritans resistance. Our results showed the complex polygenic architecture of resistance of large yellow croaker against C. irritans. These results would be helpful for the researches of the molecular mechanism of C. irritans resistance and genome-assisted breeding of large yellow croaker.

Genome-Wide Association Study Identifies Genomic Loci of Sex Determination and Gonadosomatic Index Traits in Large Yellow Croaker (Larimichthys crocea).

Larimichthys crocea is one of the traditional marine culture fishes in China, widely distributed in South China Sea, East Sea, and southern Yellow Sea. Sex dimorphism is evident in this species that females present a substantial growth strength than males, suggesting breeding females could obtain more economic benefits in L. crocea aquaculture industry. With the continuous expansion of aquaculture industry, both identifying sex-associated genome region and understanding the genetic basis underlying gonad differentiation and development matter to not only sex control aquaculture but also breeding industry. Thus, genome-wide association analysis (GWAS) of sex determination was conducted with a random breeding population of 905 individuals (including 463 females and 442 males) by ddRAD sequencing. For sex determination, 21 significant single nucleotide polymorphisms (SNPs) in chromosome (Chr) 22 were identified. Surrounding these SNPs, we founded 14 candidate genes, including dmrt1, dmrt3, and piwil2, fam102a, and odf2. The sex-associated region was narrowed down further to 2.4 Mb on Chr22 through Fst scanning and insertion-deletion (InDel) analysis. Besides, 3 SNPs in the supposed sex-determining region on Chr22 were identified as highly associated with gonad differentiation through GWAS on gonadosomatic index (GSI) in 350 males and 231 females. Because of the significant difference of GSI between females and males of L. crocea, GWAS on GSI of different genders was also conducted independently. Finally, we identified a SNP in Chr18 showing genome-wide significant association with male GSI (MGSI) and three genes axl, cyp2a10, and cyp2g1 involved in the gonadal development regulation process of aromatase. Overall, this study explored the genetic basis of sex determination mechanism and provided novel insights into gonad differentiation and development, offering solid genetic support for sex control breeding, marker-assisted selection, and marine resources conservation.

Chromosome-Level Assembly of the Southern Rock Bream (Oplegnathus fasciatus) Genome Using PacBio and Hi-C Technologies.

The Rock Bream (Oplegnathus fasciatus) is an economically important rocky reef fish of the Northwest Pacific Ocean. In recent years, it has been cultivated as an important edible fish in coastal areas of China. Despite its economic importance, genome-wide adaptions of domesticated O. fasciatus are largely unknown. Here we report a chromosome-level reference genome of female O. fasciatus (from the southern population in the subtropical region) using the PacBio single molecule sequencing technique (SMRT) and High-through chromosome conformation capture (Hi-C) technologies. The genome was assembled into 120 contigs with a total length of 732.95 Mb and a contig N50 length of 27.33 Mb. After chromosome-level scaffolding, 24 chromosomes with a total length of 723.22 Mb were constructed. Moreover, a total of 27,015 protein-coding genes and 5,880 ncRNAs were annotated in the reference genome. This reference genome of O. fasciatus will provide an important resource not only for basic ecological and population genetic studies but also for dissect artificial selection mechanisms in marine aquaculture.

Identification and Expression Analysis of Long Non-coding RNA in Large Yellow Croaker (Larimichthys crocea) in Response to Cryptocaryon irritans Infection.

Large-scale transcription studies have revealed numerous lncRNAs (long non-coding RNAs). lncRNAs have been proposed to participate in the regulation of a diverse range of biological processes, including transcriptional regulation. Although lncRNAs have attracted increasing attention, the studies in large yellow croaker (Larimichthys crocea) are still rare, and they lack systematic analysis. In this study, 101 RNA-seq datasets varied in ages, sexes, and tissues were retrieved from the NCBI database to generate a comprehensive catalog of large yellow croaker transcriptome database. A set of 14,599 high-confidence lncRNAs from 13,673 loci were identified and characterized. Furthermore, RNA-seq datasets obtained from the infection of C. irritans were employed to investigate the differential expression pattern of lncRNAs and analyze potential biological functions. A total of 77 differentially expressed lncRNAs targeting to 567 protein-coding genes were identified by using expression analysis. Several immune genes, including TLR5, CD2AP, and MMP9, were highlighted. With GO enrichment and KEGG pathway analysis, the immune-related terms or pathways were enriched. This study created a comprehensive dataset of lncRNAs for large yellow croaker, which would be helpful for the researches of functional roles of lncRNAs in large yellow croaker.

Development and Evaluation of a High-Throughput Single-Nucleotide Polymorphism Array for Large Yellow Croaker (Larimichthys crocea).

High-density single-nucleotide polymorphism (SNP) genotyping array is an essential tool for genetic analyses of animals and plants. Large yellow croaker (Larimichthys crocea) is one of the most commercially important marine fish species in China. Although plenty of SNPs have been identified in large yellow croaker, no high-throughput genotyping array is available. In this study, a high-throughput SNP array named NingXin-I with 600K SNPs was developed and evaluated. A set of 82 large yellow croakers were collected from different locations of China and re-sequenced. A total of 9.34M SNPs were identified by mapping sequence reads to the large yellow croaker reference genome. About 1.98M candidate SNPs were selected for further analyses by using criteria such as SNP quality score and conversion performance in the final array. Finally, 579.5K SNPs evenly distributed across the large yellow croaker genome with an average spacing of 1.19 kb were proceeded to array production. The performance of NingXin-I array was evaluated in 96 large yellow croaker individuals from five populations, and 83.38% SNPs on the array were polymorphic sites. A further test of the NingXin-I array in five closely related species in Sciaenidae identified 26.68-56.23% polymorphic SNP rate across species. A phylogenetic tree inferred by using the genotype data generated by NingXin-I confirmed the phylogenetic distance of the species in Sciaenidae. The performance of NingXin-I in large yellow croaker and the other species in Sciaenidae suggested high accuracy and broad application. The NingXin-I array should be valuable for quantitative genetic studies, such as genome-wide association studies (GWASs), high-density linkage map construction, haplotype analysis, and genome-based selection.