Whole-genome resequencing provides insights into the population structure and domestication signatures of ducks in eastern China.

BACKGROUND: Duck is an ancient domesticated animal with high economic value, used for its meat, eggs, and feathers. However, the origin of indigenous Chinese ducks remains elusive. To address this question, we performed whole-genome resequencing to first explore the genetic relationship among variants of these domestic ducks with their potential wild ancestors in eastern China, as well as understand how the their genomes were shaped by different natural and artificial selective pressures. RESULTS: Here, we report the resequencing of 60 ducks from Chinese spot-billed ducks (Anas zonorhyncha), mallards (Anas platyrhnchos), Fenghua ducks, Shaoxing ducks, Shanma ducks and Cherry Valley Pekin ducks of eastern China (ten from each population) at an average effective sequencing depth of ~ 6× per individual. The results of population and demographic analysis revealed a deep phylogenetic split between wild (Chinese spot-billed ducks and mallards) and domestic ducks. By applying selective sweep analysis, we identified that several candidate genes, important pathways and GO categories associated with artificial selection were functionally related to cellular adhesion, type 2 diabetes, lipid metabolism, the cell cycle, liver cell proliferation, and muscle functioning in domestic ducks. CONCLUSION: Genetic structure analysis showed a close genetic relationship of Chinese spot-billed ducks and mallards, which supported that Chinese spot-billed ducks contributed to the breeding of domestic ducks. During the long history of artificial selection, domestic ducks have developed a complex biological adaptation to captivity.

The circular RNA aplacirc_13267 upregulates duck granulosa cell apoptosis by the apla-miR-1-13/THBS1 signaling pathway.

Follicle development is a key factor that determines the reproductive performance of poultry. The existing evidence suggests that circular RNAs (circRNAs) play an important role in a variety of biological processes, especially in posttranscriptional regulation, but the regulatory mechanism of circRNAs in duck follicle development has rarely been reported. To better explore the molecular mechanism of follicle development in ducks, we sequenced and analyzed the follicular circRNAs; 4,204 circRNAs were predicted in the duck follicles. Fourteen circRNAs were differentially expressed between the white follicles and yellow follicles. The results of our studies showed that aplacirc_013267 promoted cell apoptosis in duck GCs. Moreover, a bioinformatics prediction analysis demonstrated that aplacirc_013267 was involved in a circRNA-miRNA-mRNA coexpression network and was observed to sponge two follicle-related miRNAs by a luciferase activity assay. Furthermore, we found that overexpression of aplacirc_013267 significantly increased thrombospondin-1 (THBS1) expression and downregulated granulosa cell apoptosis. The mechanistic study showed that aplacirc_013267 directly binds to and inhibits apla-mir-1-13; then, aplacirc_013267 increases the expression of THBS1 and upregulates granulosa cell apoptosis. Taken together, our findings demonstrate that circRNAs have potential effects in duck ovarian follicles and that circRNAs may represent a new avenue to understand follicular development.

Comparative transcriptome analysis reveals PERP upregulated during Salmonella Enteritidis challenge in laying ducks.

Salmonella Enteritidis (SE) can be transmitted to eggs through cecum or the ovary from infected layers and causes food poisoning in humans. The mechanism of cecal transmission has been extensively studied. However, the mechanism and route of transovarian transmission of SE remain unclear. In this study, the ducks were orally inoculated with SE, and the ovarian follicles and stroma were collected to detect SE infection. The immune responses were triggered and the innate and adaptive immune genes (TLR4, NOD1, AvßD7, and IL-1ß) were upregulated significantly during the SE challenge. Moreover, the ovary tissues (small follicle and stroma) of susceptible and resistant-laying ducks were performed by RNA sequencing. We obtained and identified 23 differentially expressed genes (DEGs) between susceptible and resistant-laying ducks in both small follicle and stroma tissues ( p < 0.05). The DEGs were predominately identified in the p53 signaling pathway. The expression of key genes (p53, MDM2, PERP, caspase-3, and Bcl-2) involved in the signaling pathway was significantly higher in granulosa cells (dGCs) from SE-infected ducks than those from uninfected ducks. Moreover, the overexpression of PERP resulted in further induction of p53, MDM2, caspase-3, and Bcl-2 during SE infection in dGCs. Whereas, an opposite trend was observed with the knockdown of PERP. Besides, it is further revealed that the PERP could enhance cell apoptosis, SE adhesion, and SE invasion in SE-infected dGCs overexpression. Altogether, our results demonstrate the duck PERP involved in the ovarian local immune niche through p53 signaling pathway in dGCs challenged with SE.

Purification of therapeutic proteins mediated by in vivo polyester immobilized sortase.

OBJECTIVES: To overcome laborious and costly procedures often associated with therapeutic protein production and purification, in vivo polyester immobilized sortase is explored for the production of human tumor necrosis factor alpha (TNFα) and human interferon alpha 2b (IFNα2b) by Escherichia coli. RESULTS: Hybrid genes encoding PhaC-Sortase-TNFα or PhaC-Sortase-IFNα2b fusions (with a LPETG recognition signal immediately before TNFα or IFNα2b), mediated intracellular production of polyester (polyhydroxyalkanoate, PHA) beads in Escherichia coli. Upon isolation of respective PHA beads, pure soluble TNFα or IFNα2b was released by activating sortase via addition of CaCl2 and triglycine. TNFα and IFNα2b each were recognized by corresponding conformational antibodies in an ELISA assay. CONCLUSIONS: In vivo polyester immobilized sortase could be exploited for production and purification of high-value therapeutic proteins without laborious and costly downstream processing.

Purification of target proteins from intracellular inclusions mediated by intein cleavable polyhydroxyalkanoate synthase fusions.

BACKGROUND: Recombinant protein production and purification from Escherichia coli is often accompanied with expensive and complicated procedures, especially for therapeutic proteins. Here it was demonstrated that, by using an intein cleavable polyhydroxyalkanoate synthase fusion, recombinant proteins can be first produced and sequestered on a natural resin, the polyhydroxyalkanoate (PHA) inclusions, then separated from contaminating host proteins via simple PHA bead isolation steps, and finally purified by specific release into the soluble fraction induced by a pH reduction. RESULTS: By translationally fusing a target protein to PHA synthase using a self-cleaving intein as linker, intracellular production of PHA beads was achieved. Upon isolation of respective PHA beads the soluble pure target protein was released by a simple pH shift to 6. The utility of this approach was exemplified by producing six target proteins, including Aequorea victoria green fluorescent protein (GFP), Mycobacterium tuberculosis vaccine candidate Rv1626, the immunoglobulin G (IgG) binding ZZ domain of protein A derived from Staphylococcus aureus, human tumor necrosis factor alpha (TNFα), human granulocyte colony-stimulating factor (G-CSF), and human interferon alpha 2b (IFNα2b). CONCLUSIONS: Here a new method for production and purification of a tag-less protein was developed through intein cleavable polyhydroxyalkanoate synthase fusion. Pure target protein could be easily obtained without laborious downstream processing.

[Fast-onset antidepressant potentials of essential oil of herbs].

The existing antidepressants demonstrated delayed onset of clinical effects, so fast-onset antidepressants are required. Essential oil of herbs showed potentials fast-onset antidepressant potential. First, its aromatic odor can directly activate olfactory nerves; its high lipophilicity causes a high blood-brain barrier penetration rate; and its high volatility is suitable for nasal-brain targeting and inhalation delivery. Therefore, essential oils can rapidly regulate brain functions by multiple ways, suggesting a fast-onset antidepressant potential. Second, the advance of studies on chemistry and pharmacology of antidepressant essential oils demonstrated chemical substances, antidepressant effects and possible action mechanisms of antidepressant essential oils. Third, the effect of essential oils' antidepressant components on fast-onset antidepressant targets was investigated. It was found that chemical constituents of essential oils antagonized NMDA receptor activities, suggesting that essential oils have fast-onset antidepressant effect. Finally, characteristics of essential oils, fast-onset antidepressant targets and drug delivery methods are integrated to give full play to essential oils' fast-onset antidepressant advantage and provide a new direction for new drug discovery.

Bioengineering of bacteria to assemble custom-made polyester affinity resins.

Proof of concept for the in vivo bacterial production of a polyester resin displaying various customizable affinity protein binding domains is provided. This was achieved by engineering various protein binding domains into a bacterial polyester-synthesizing enzyme. Affinity binding domains based on various structural folds and derived from molecular libraries were used to demonstrate the potential of this technique. Designed ankyrin repeat proteins (DARPins), engineered OB-fold domains (OBodies), and VHH domains from camelid antibodies (nanobodies) were employed. The respective resins were produced in a single bacterial fermentation step, and a simple purification protocol was developed. Purified resins were suitable for most lab-scale affinity chromatography purposes. All of the affinity domains tested produced polyester beads with specific affinity for the target protein. The binding capacity of these affinity resins ranged from 90 to 600 nmol of protein per wet gram of polyester affinity resin, enabling purification of a recombinant protein target from a complex bacterial cell lysate up to a purity level of 96% in one step. The polyester resin was efficiently produced by conventional lab-scale shake flask fermentation, resulting in bacteria accumulating up to 55% of their cellular dry weight as polyester. A further proof of concept demonstrating the practicality of this technique was obtained through the intracellular coproduction of a specific affinity resin and its target. This enables in vivo binding and purification of the coproduced "target protein." Overall, this study provides evidence for the use of molecular engineering of polyester synthases toward the microbial production of specific bioseparation resins implementing previously selected binding domains.

In vivo polyester immobilized sortase for tagless protein purification.

BACKGROUND: Laboratory scale recombinant protein production and purification techniques are often complicated, involving multiple chromatography steps and specialized equipment and reagents. Here it was demonstrated that recombinant proteins can be expressed as covalently immobilized to the surface of polyester (polyhydroxyalkanoate, PHA) beads in vivo in Escherichia coli by genetically fusing them to a polyester synthase gene (phaC). The insertion of a self-cleaving module, a modified sortase A (SrtA) from Staphylococcus aureus and its five amino acid recognition sequence between the synthase and the target protein led to a simple protein production and purification method. RESULTS: The generation of hybrid genes encoding tripartite PhaC-SrtA-Target fusion proteins, enabled immobilization of proteins of interest to the surface of PHA beads in vivo. After simple cell lysis and isolation of the PHA beads, the target proteins could be selectively and efficiently released form the beads by activating the sortase with CaCl2 and triglycine. Up to 6 mg/l of soluble proteins at a purity of ~98 % could be isolated in one step with no optimization. This process was used to produce and isolate three proteins: Green fluorescent protein, maltose binding protein and the Mycobacterium tuberculosis vaccine candidate Rv1626. CONCLUSIONS: We have developed a new technique for easy production and purification of recombinant proteins. This technique is capable of producing and purifying high yields of proteins suitable for research application in less than 2 days. No costly or specialized protein chromatography equipment, resins, reagents or expertise are required.

Polyhydroyxalkanoate synthase fusions as a strategy for oriented enzyme immobilisation.

Polyhydroxyalkanoate (PHA) is a carbon storage polymer produced by certain bacteria in unbalanced nutrient conditions. The PHA forms spherical inclusions surrounded by granule associate proteins including the PHA synthase (PhaC). Recently, the intracellular formation of PHA granules with covalently attached synthase from Ralstonia eutropha has been exploited as a novel strategy for oriented enzyme immobilisation. Fusing the enzyme of interest to PHA synthase results in a bifunctional protein able to produce PHA granules and immobilise the active enzyme of choice to the granule surface. Functionalised PHA granules can be isolated from the bacterial hosts, such as Escherichia coli, and maintain enzymatic activity in a wide variety of assay conditions. This approach to oriented enzyme immobilisation has produced higher enzyme activities and product levels than non-oriented immobilisation techniques such as protein inclusion based particles. Here, enzyme immobilisation via PHA synthase fusion is reviewed in terms of the genetic designs, the choices of enzymes, the control of enzyme orientations, as well as their current and potential applications.

Genome-Wide Association Study of Egg Production Traits in Shuanglian Chickens Using Whole Genome Sequencing.

Egg production is the most important economic trait in laying hens. To identify molecular markers and candidate genes associated with egg production traits, such as age at first egg (AFE), weight at first egg (WFE), egg weight (EW), egg number (EN), and maximum consecutive egg laying days (MCD), a genome-wide analysis by whole genome sequencing was performed in Shuanglian chickens. Through whole genome sequencing and quality control, a total of 11,006,178 SNPs were obtained for further analysis. Heritability estimates ranged from moderate to high for EW (0.897) and MCD (0.632), and from low to moderate (0.193~0.379) for AFE, WFE, and EN. The GWAS results showed 11 genome-wide significant SNPs and 23 suggestive significant SNPs were identified to be associated with EN, MCD, WFE, and EW. Linkage disequilibrium analysis revealed twenty-seven SNPs associated with EN were located in a 0.57 Mb region on GGA10, and clustered into five blocks. Through functional annotation, three candidate genes NEO1, ADPGK, and CYP11A1, were identified to be associated with EN, while the S1PR4, LDB2, and GRM8 genes was linked to MCD, WFE, and EW, respectively. These findings may help us to better understand the molecular mechanisms underlying egg production traits in chickens and contribute to genetic improvement of these traits.

Characteristics and removal of antibiotics and antibiotic resistance genes in a constructed wetland from a drinking water source in the Yangtze River Delta.

Safe drinking water is crucial to public health. However, approximately one-third of the world's population lacks access to clean drinking water. The presence of antibiotics and antibiotic resistance genes (ARGs) in drinking water sources has become a severe problem worldwide due to its potential threat to human health. We monitored the occurrence and variations of 23 antibiotics and 9 ARGs in different treatment processes in a constructed wetland serving as drinking water source in the Yangtze River Delta, China. The studied wetland is consisted of four treatment processes: pretreatment area, pump station lifting, root-channel ecological purification area and deep purification area. Except for sulfapyridine and roxithromycin, 21 antibiotics were detected at concentrations ranging from 0.15 to 59.52 ng/L. The concentration of macrolides was the highest in this wetland, especially tylosin (42.86-59.52 ng/L). TetG, tetX and sul2 were the dominant ARGs in both water (2.41 × 10-4-1.87 × 10-2) and sediment (6.65 × 10-5-4.92 × 10-3). In addition, a strong correlation between ARGs in water and ARGs in sediment (Pearson, R2 > 0.9, p < 0.05) indicated an exchange between the two phases. Moreover, the significantly positive correlation of ARGs between the inlet and outlet of each subsystem illustrated that upstream pollution was the primary source for downstream processes. In general, the wetland system could efficiently eliminate antibiotics (9.0-53.8%) and ARGs (14.5-94.1%), with the deep purification area having the highest removal efficiency. Overall, our results provide important insights into the occurrence, abundance and removal of antibiotics and ARGs in the constructed wetland serving as drinking water sources.

Effects of Peroxyl Radicals on the Structural Characteristics and Fatty Acid Composition of High-Density Lipoprotein from Duck Egg Yolk.

In this study, high-density lipoprotein (HDL) from duck egg yolk was subjected to oxidation with a system based on 2,2'-azobis (2-amidinopropane) dihydrochloride (AAPH)-derived peroxyl radicals. The effects of peroxyl radicals on the protein carbonyl, free sulfhydryl, secondary/tertiary structure, surface hydrophobicity, solubility, particle size distribution, zeta potential and fatty acid composition of HDL were investigated by using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), Fourier-transform infrared spectroscopy (FTIR), circular dichroism (CD), fluorescence spectroscopy, dynamic light scattering and ultra-high-performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS). The results indicated that the content of protein carbonyl was significantly increased, that of free sulfhydryl was obviously reduced, and the ordered secondary structure was also decreased with increasing AAPH concentration. In addition, the surface hydrophobicity and solubility of HDL showed apparent increases due to the exposure of hydrophobic groups and aggregation of protein caused by oxidation. The fatty acid composition of HDL exhibited pronounced changes due to the disrupted protein-lipid interaction and lipid oxidation by AAPH-derived peroxyl radicals. These results may help to elucidate the molecular mechanism for the effect of lipid oxidation products on the oxidation of duck yolk proteins.

Multiomic analysis revealed the regulatory role of the KRT14 gene in eggshell quality.

Background: Eggshell strength and thickness are critical factors in reducing the egg breaking rate and preventing economic losses. The calcite biomineralization process is very important for eggshell quality. Therefore, we employed transcriptional sequencing and proteomics to investigate the differences between the uteruses of laying hens with high- and low-breaking-strength shells. Results: A total of 1,028 differentially expressed genes (DEGs) and 270 differentially expressed proteins (DEPs) were identified. The analysis results of GO terms and KEGG pathways showed that most of the DEGs and DEPs were enriched in vital pathways related to processes such as calcium metabolism, hormone and amino acid biosynthesis, and cell proliferation and apoptosis. Several DEGs and DEPs that were coexpressed at mRNA and protein levels were verified. KRT14 (keratin-14) is a candidate gene (protein) obtained by multiple omics analysis due to the fold difference of KRT14 being the largest. After the overexpression of KRT14 in uterine epithelial cells, the expressions of OC116 (ovocleididin-116), CALB1 (calbindin 1), and BST1 (ADP-ribosyl cyclase 2) were found to be increased significantly, while the expression of OC17 (ovocleididin-17) was found to be decreased significantly. Conclusion: In summary, this study confirms that during normal calcification, there are differences in ion transport between the uterus of hens producing high-breaking-strength eggshells and those producing low-breaking-strength eggshells, which may help elucidate the eggshell calcification process. The KRT14 gene may promote calcium metabolism and deposition of calcium carbonate in eggshells.

LncRNA lnc_13814 promotes the cells apoptosis in granulosa cells of duck by acting as apla-miR-145-4 sponge.

Follicle development is a vital factor which determines the reproductive performance of poultry. Long noncoding RNAs (lncRNAs) have been reported to maintain animal reproductive function and play key roles in ovarian development and hormone secretion. But the regulatory mechanism of lncRNAs in duck follicle development has seldom been reported. In this study, to better explore the molecular mechanism of follicle development in ducks, the follicular lncRNA was sequenced and analyzed. A total of 9,551 lncRNAs were predicted in the duck follicles. Four hundred and forty-five lncRNAs were differentially expressed between the white follicles and yellow follicles. The results of our studies showed that lnc_13814 promoted cell apoptosis in duck GCs. Furthermore, the bioinformatics analysis results demonstrated that lnc_13814 was involved in a lncRNA-miRNA-mRNA coexpression network and it was observed to sponge two follicle-related miRNAs by a luciferase activity assay. Moreover, we found that overexpression of lnc_13814 significantly increased DNA damage inducible transcript 3 (DDIT3) expression and downregulated GCs apoptosis. Finally, we found that lnc_13814 directly binds to and inhibits apla-mir-145-4; then, lnc_13814 increases the expression of DDIT3 and up-regulates GCs apoptosis. Taken together, our findings demonstrate that lncRNAs have potential effects on duck ovarian follicles and lncRNAs may represent a new approach to understand follicular development.

Analysis of miRNAs and their target genes associated with mucosal damage caused by transport stress in the mallard duck intestine.

Bowel health is an important factor for duck rearing that has been linked to feed uptake and growth and death rates. Because the regulatory networks associated with acute stress-mediated injury in the duck gastrointestinal tract have not clearly elucidated, we aimed to explore potential miRNA-mRNA pairs and their regulatory roles in oxidative stress injury caused by transport stress. Here, 1-day-old mallard ducklings from the same breeder flock were collected and transported for 8 h, whereas the control group was not being transported. Various parameters reflecting oxidative stress and the tissue appearance of the intestine were assessed. The data showed that the plasma T-AOC and SOD concentrations were decreased in the transported ducklings. The intestine of the transported ducklings also displayed significant damage. High-throughput sequencing of the intestine revealed 44 differentially expressed miRNAs and 75 differentially expressed genes, which constituted 344 miRNA-mRNA pairs. KEGG pathway analysis revealed that the metabolic, FoxO signaling, influenza A and TGF-ß signaling pathways were mainly involved in the mechanism underlying the induction of intestinal damage induced by simulated transport stress in ducks. A miRNA-mRNA pair, miR-217-5p/CHRDL1, was selected to validate the miRNA-mRNA negative relationship, and the results showed that miR-217-5p could influence CHRDL1 expression. This study provides new useful information for future research on the regulatory network associated with mucosal damage in the duck intestine.

Selective adipogenic effects of propionate on bovine intramuscular and subcutaneous preadipocytes.

This study investigated the regulation by propionate of proliferation and differentiation of bovine intramuscular preadipocytes (BIPs) and subcutaneous preadipocytes (BSPs) and compared the adipogenic potential of these two preadipocytes from different sites. Propionate had no significant proliferative effects on BIPs or BSPs but the proliferative potential was higher in BSPs than in BIPs in all treatment groups (P<0.05). Furthermore, propionate significantly stimulated differentiation in a dose-dependent manner in BIPs (1, 3, and 6mM, P<0.05), but did not differ from each other (P<0.05). However, propionate significantly promoted differentiation in BSPs only in the 1mM group (P<0.05). Additionally, differential effects of propionate on the differentiation of BIPs and BSPs were detected. Taken together, these results indicate that propionate has selective adipogenic effects on BIPs and BSPs, and contributes to the hypertrophy of adipocytes within intramuscular adipose tissue but to the recruitment of new adipocytes within subcutaneous adipose tissue.

EGFR promotes the proliferation of quail follicular granulosa cells through the MAPK/extracellular signal-regulated kinase (ERK) signaling pathway.

Follicles develop into preovulatory follicles during folliculogenesis and the majority of small yellow follicles become atretic and gets reabsorbed. In this study, based the RNA-seq results of duck ovary, epidermal growth factor receptor (EGFR) was selected as a candidate gene in follicular development and the role was explored. The results demonstrated that EGFR-P8 was the quail EGFR core promoter. It had an E2F4 binding site within EGFR core promoter. E2F4 overexpression significantly increased EGFR expression in quail granulosa cells (GCs). However, the effect was abolished when the GCs were treated with corynoxeine, an inhibitor of the mitogen-activated protein kinase/extracellular regulated protein kinase (MAPK/ERK) signaling pathway. Moreover, luciferase reporter assay and chromatin immunoprecipitation experiments showed that E2F4 upregulated the expression of EGFR expression, which increased E2 and P4 production. In addition, EGFR regulated GCs proliferation and affected follicular development. Taken together, our findings suggested that EGFR, which was regulated by E2F4, enhanced the expression of MAPK/ERK pathway components and follicular development. These results provided an important basis for an improved understanding of the MAPK/ERK pathway and new insight into the development of quail follicles.

Dynamic Expression Pattern of SERPINA1 Gene from Duck (Anas platyrhynchos).

SERPINA1 is a member of serine protease inhibitors and is increasingly considered to be a regulator of innate immunity in human and animals. However, the expression and function of SERPINA1 gene in immune defense against viral infection remain unknown in ducks. The full-length du SERPINA1 cDNA sequence was obtained using reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). It contained 1457 nucleotide, including 47-bp 5' UTR, 135-bp 3' UTR, and 1275-bp open reading frame (ORF), and encodes a 424-amino acid protein. Then, the tissue expression profile of du SERPINA1 gene was determined. Real-time quantitative polymerase chain reaction (real-time qPCR) analysis revealed that du SERPINA1 mRNA is ubiquitous in various tissues, but higher expression levels were observed in lung and liver tissues. In addition, the expression pattern was investigated when the ducklings were challenged with duck hepatitis virus 1(DHV-1) and polyriboinosinic polyribocytidylic acid (poly I:C). After DHV-1 injection or poly I:C treatment, du SERPINA1 mRNA was up-regulated in the liver and kidney tissues. However, the peak time in two tissues was not consistent. In kidney, the expression lever of SERPINA1 increased immediately after the treatment while in liver tissue it kept steady until 12 h post-infection. Our results indicate that SERPINA1 has an active role in the antiviral response, and thus improve our understanding of the role of this protein.

Population genomic data reveal genes related to important traits of quail.

Background: Japanese quail (Coturnix japonica), a recently domesticated poultry species, is important not only as an agricultural product, but also as a model bird species for genetic research. However, most of the biological questions concerning genomics, phylogenetics, and genetics of some important economic traits have not been answered. It is thus necessary to complete a high-quality genome sequence as well as a series of comparative genomics, evolution, and functional studies. Results: Here, we present a quail genome assembly spanning 1.04 Gb with 86.63% of sequences anchored to 30 chromosomes (28 autosomes and 2 sex chromosomes Z/W). Our genomic data have resolved the long-term debate of phylogeny among Perdicinae (Japanese quail), Meleagridinae (turkey), and Phasianinae (chicken). Comparative genomics and functional genomic data found that four candidate genes involved in early maturation had experienced positive selection, and one of them encodes follicle stimulating hormone beta (FSHß), which is correlated with different FSHß levels in quail and chicken. We re-sequenced 31 quails (10 wild, 11 egg-type, and 10 meat-type) and identified 18 and 26 candidate selective sweep regions in the egg-type and meat-type lines, respectively. That only one of them is shared between egg-type and meat-type lines suggests that they were subject to an independent selection. We also detected a haplotype on chromosome Z, which was closely linked with maroon/yellow plumage in quail using population resequencing and a genome-wide association study. This haplotype block will be useful for quail breeding programs. Conclusions: This study provided a high-quality quail reference genome, identified quail-specific genes, and resolved quail phylogeny. We have identified genes related to quail early maturation and a marker for plumage color, which is significant for quail breeding. These results will facilitate biological discovery in quails and help us elucidate the evolutionary processes within the Phasianidae family.

Association of VIPR-1 gene polymorphisms and haplotypes with egg production in laying quails.

The laying quail is a worldwide breed which exhibits high economic value. In our current study, the vasoactive intestinal peptide receptor-1 (VIPR-1) was selected as the candidate gene for identifying traits of egg production. A single nucleotide polymorphism (SNP) detection was performed in 443 individual quails, including 196 quails from the H line, 202 quails from the L line, and 45 wild quails. The SNPs were genotyped using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Two mutations (G373T, A313G) were detected in all the tested quail populations. The associated analysis showed that the SNP genotypes of the VIPR-1 gene were significantly linked with the egg weight of G373T and A313G in 398 quails. The quails with the genotype GG always exhibited the largest egg weight for the two mutations in the H and L lines. Linkage disequilibrium (LD) analysis indicated that G373T and A313G loci showed the weakest LD. Seven main diplotypes from the four main reconstructed haplotypes were observed, indicating a significant association of diplotypes with egg weight. Quails with the h1h2 (GGGT) diplotype always exhibited the smallest egg weight and largest egg number at 20 weeks of age. The overall results suggest that the alterations in quails may be linked with potential major loci or genes affecting reproductive traits.