Molecular characterization, expression profile and transcriptional regulation of the CYP19 gene in goose ovarian follicles.

Cytochrome P450 Family 19 (CYP19) is a crucial enzyme to catalyze the conversion of androgens to estrogens. However, the regulatory mechanism of goose CYP19 gene remains poorly understood. The present study attempted to obtain the full-length coding sequence (CDS) and 5'-flanking sequence of CYP19 gene, to investigate its expression and distribution profiles in different sized follicles, and to analyze the transcriptional regulatory mechanism of CYP19 gene in goose. Results showed that its CDS consisted of 1512 nucleotides and the encoded amino acid sequence contained a classical P450 structural domain. Homology analysis showed that there were high homologies of nucleotide and amino acid sequences between goose and other avian species. Its promoter sequence spanned from -1925 bp to the transcription start site (ATG) and several transcriptional factors were predicted in this region. Further analysis from luciferase assay showed that the luciferase activity was the highest spanning from -118 to -1 bp by constructing deletion promoter reporter vector. In addition, result from quantitative real-time polymerase chain reaction indicated that the mRNA level of CYP19 gene were highly expressed in theca layer of the fifth largest follicle, and the cellular location was in the theca externa cells by immunohistochemistry. Taken together, it could be concluded that the transcription activity of CYP19 gene was activated by transcriptional factors in its proximal region of promoter to promote the synthesis of estrogens, regulating the selection of pre-hierarchical into hierarchical follicle in goose.

Identifying molecular pathways and candidate genes associated with knob traits by transcriptome analysis in the goose (Anser cygnoides).

Anser cygnoides has a spherical crest on the beak roof, which is described as knob. However, the mechanisms affecting knob morphology are unclear. Here, we investigated the phenotypic characteristics and molecular basis of knob-size differences in Yangzhou geese. Anatomically, the knob was identified as frontal hump in the frontal area of the skull, rather than hump of upper beak. Although the frontal hump length, and height varied greatly in geese with different knob phenotypes, little was changed in the width. Histologically, knob skin in large-size knobs geese have a greater length in the stratum corneum, stratum spinosum, and stratum reticular than that in small-size knobs geese. Moveover, the 415 differentially expressed genes were found between the large knobs and small ones through transcriptome profiling. In addition, GO enrichment and KEGG pathway analysis revealed 455 significant GO terms and 210 KEGG pathways were enriched, respectively. Among these, TGF-ß signaling and thyroid hormone synthesis-signaling pathways were identified to determine knob-size phenotype. Furthermore, BMP5, DCN, TSHR and ADCY3 were recognized to involve in the growth and development of knob. Our data provide comprehensive molecular determinants of knob size phenotype, which can potentially promote the genetic improvement of goose knobs.

Geometrical characteristics of eggs from 3 poultry species.

We studied the correlations between egg geometrical parameters (i.e., egg shape index, sphericity, geometric mean diameter, surface area, and volume) and eggshell qualities, or the organic matrix in eggshell. Eggs were collected from 5 poultry breeds belonging to 3 species (commercial Hy-line Brown Chicken, Shaoxing Duck, Jinding Duck, Taihu Goose, and Zhedong White Goose). The geometrical parameters showed high variation among 3 species of poultry, and even between breeds in the same species. The five geometrical parameters were grouped into 2 sets, one contained shape index and sphericity, the other comprised geometric mean diameter, surface area, and volume. The parameters in the same set can be perfectly fitted to one another. Egg weight, shell membrane weight, and calcified shell weight were significantly correlated with geometric mean diameter, surface area, and volume. In accordance with false discovery rate-adjusted P value, both shell membrane relative weight and calcified shell thickness showed no significant correlations with any of the geometrical parameters. However, the correlations between geometrical parameters and other shell variables (calcified shell weight, shell relative weight, calcified shell thickness uniformity, and eggshell breaking strength) depend on breed. Both constitutive proportions and percentage contents of 3 eggshell matrix components (acid-insoluble, water-insoluble, and both acid and water facultative-soluble matrix) had no effects on egg shape and size. The correlations between the amounts of various shell matrix, egg shape and size depend on breed or species. This study provides a methodology and the correlation between geometrical parameters and eggshell qualities, and between geometrical parameters and organic matrix components in calcified shells.

The protein and fat quality of thigh muscles from Polish goose varieties.

This study aimed to evaluate the nutritional value of thigh meat from 4 Polish geese varieties. Protein, fat, and cholesterol content, as well as amino acid and fatty acid profiles, were determined. Based on the percentage of amino acid in protein and fatty acids in meat lipids, the health lipid indices were calculated. The experimental material covered thigh muscles from 17-week-old Kartuska (Ka), Suwalska (Su), Lubelska (Lu), and Kielecka (Ki) geese reared in a semi-intensive system. Muscle protein content did not differ significantly between varieties. The protein content of the Ka, Su, Lu, and Ki goose meat was deemed high-value as it contained all the essential amino acids in the proportions consistent with standard protein values. The muscles of all the researched geese varieties were characterized by a high level of Lys, which indicates that this meat is a good source of it (AASLys 240-280%). Current findings showed that polyunsaturated fatty acids (PUFA)/saturated fatty acids (SFA) and PUFA n-6/n-3 ratios in Ka, Su, Lu, and Ki muscles were found to be within the optimum values for human diets. No significant differences were observed in monounsaturated fatty acids, PUFA, and unsaturated fatty acids (UFA) between the analyzed muscles. The meat of Ka and Su contained significantly more lipids than Lu and Ki. A more beneficial amino acid profile was found in Ka meat due to a higher content of PUFA n-3 and the best n-6/n-3 ratio in comparison with other varieties. The muscles of the Ka variety also contained the least cholesterol. However, the Ki goose muscles stood out among other varieties with the least percentage of SFA, the highest share of docosahexaenoic acid (C 22:6 n-3), as well as the most beneficial value of the following indices: UFA/SFA, hypocholesterolemic fatty acid/hypercholesterolemic fatty acid ratio, and nutritive value index. The thigh muscles of Ka, Su, Lu, and Ki were characterized by an atherogenicity index that met the levels of recommended values (<1) in the diet of a human being, while the thrombogenicity index was slightly higher than the recommended value (<0.5).

Recent introgression between Taiga Bean Goose and Tundra Bean Goose results in a largely homogeneous landscape of genetic differentiation.

Several studies have uncovered a highly heterogeneous landscape of genetic differentiation across the genomes of closely related species. Specifically, genetic differentiation is often concentrated in particular genomic regions ("islands of differentiation") that might contain barrier loci contributing to reproductive isolation, whereas the rest of the genome is homogenized by introgression. Alternatively, linked selection can produce differentiation islands in allopatry without introgression. We explored the influence of introgression on the landscape of genetic differentiation in two hybridizing goose taxa: the Taiga Bean Goose (Anser fabalis) and the Tundra Bean Goose (A. serrirostris). We re-sequenced the whole genomes of 18 individuals (9 of each taxon) and, using a combination of population genomic summary statistics and demographic modeling, we reconstructed the evolutionary history of these birds. Next, we quantified the impact of introgression on the build-up and maintenance of genetic differentiation. We found evidence for a scenario of allopatric divergence (about 2.5 million years ago) followed by recent secondary contact (about 60,000 years ago). Subsequent introgression events led to high levels of gene flow, mainly from the Tundra Bean Goose into the Taiga Bean Goose. This scenario resulted in a largely undifferentiated genomic landscape (genome-wide FST = 0.033) with a few notable differentiation peaks that were scattered across chromosomes. The summary statistics indicated that some peaks might contain barrier loci while others arose in allopatry through linked selection. Finally, based on the low genetic differentiation, considerable morphological variation and incomplete reproductive isolation, we argue that the Taiga and the Tundra Bean Goose should be treated as subspecies.

Molecular cloning, characterisation, and expression analysis of adipocyte fatty acid binding protein gene in Xupu goose (Anser cygnoides domesticus).

1. Adipocyte fatty acid binding protein (A-FABP) plays a key role in fatty acid uptake and intracellular transport. The objective of the present study was to identify and characterise the A-FABP gene in Xupu goose.2. The full-length cDNA of goose A-FABP gene was cloned from the liver tissue using reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). The distribution of the goose A-FABP in different tissues was determined by quantitative real-time PCR (qRT-PCR).3. The results showed that the full-length cDNA sequence of goose A-FABP was 657 bp, containing a 5'-UTR of 52 bp, a 3'-UTR of 206 bp and an open reading frame (ORF) of 399 bp, which encoded a polypeptide of 132 amino acids (AA).4. The AA sequence of goose A-FABP showed 76.52%, 75.00%, 93.18% and 99.24% identities with previously described homologues from humans (Homo sapiens), mouse (Mus musculus), chicken (Gallus gallus), and duck (Anas platyrhynchos), respectively, and phylogenetic analysis revealed a close relationship among them. The transcript of Xupu goose A-FABP was ubiquitously expressed in all tested tissues, and showed a high-level expression in abdominal fat, sebum and liver.5. A significant positive correlation was identified between A-FABP mRNA abundance in the three adipose tissues and liver weight, ratio of liver to body weight, TG content, and VLDL concentration in the plasma of Xupu goose. A significant negative correlation was observed between the mRNA level of A-FABP and HDL concentration in the plasma of Xupu goose.6. These findings provide a foundation for further research on the function and mechanism of A-FABP in the fat deposition process.

Crucial sites and environmental variables for wintering migratory waterbird population distributions in the natural wetlands in East Dongting Lake, China.

Dongting Lake is the second largest freshwater lake in China and is one of the globally important wintering sites for migratory waterbirds in the East Asian-Australasian Flyway. Crucial sites and environmental variables for wintering migratory waterbirds are of great concern in the Dongting Lake wetlands. In this research, based on annual (2003/2004-2016/2017) waterbird and habitat census data, we recognized the crucial sites for waterbirds during wintering seasons by comparing the difference of waterbird populations at the community, foraging guild and species levels in different natural wetlands within East Dongting Lake, and then identified the crucial environmental variables affecting waterbird distributions by analyzing the relationship between waterbird populations and the environmental variables, including vegetation area, mudflat area, water area with the depth of 0-20 cm, water area with the depth of 20-50 cm, water area with the depth of 50-100 cm, water area with the depth >100 cm, growth status of vegetation (Min, Mean and Max NDVI), and the human disturbance. Results indicated that five natural wetlands, i.e., Daxiaoxi, Chunfeng, Baihu lakes, Dingzi dyke and Tanjiaweizi, were recognized as the crucial wintering sites for migratory waterbirds in the East Dongting Lake. Among the ten selected environmental variables, water areas with the depth of 0-20 cm, 20-50 cm and >100 cm, human disturbance, Min and Mean NDVIs were identified as the crucial environmental variables overall. Waterbirds at different levels exhibited significant linear relationship with certain environmental variables, with the exception of Bean goose and Lesser White-fronted goose at the species level, which showed Gaussian distribution with changes in mean NDVI. The crucial environmental variables appeared to be foraging guild- and species-specific. These findings provide significant information for managers to understand the differences of wetlands and waterbird populations within East Dongting Lake, and to make more targeted conservation efforts.

Molecular basis of hemoglobin adaptation in the high-flying bar-headed goose.

During the adaptive evolution of a particular trait, some selectively fixed mutations may be directly causative and others may be purely compensatory. The relative contribution of these two classes of mutation to adaptive phenotypic evolution depends on the form and prevalence of mutational pleiotropy. To investigate the nature of adaptive substitutions and their pleiotropic effects, we used a protein engineering approach to characterize the molecular basis of hemoglobin (Hb) adaptation in the high-flying bar-headed goose (Anser indicus), a hypoxia-tolerant species renowned for its trans-Himalayan migratory flights. To test the effects of observed substitutions on evolutionarily relevant genetic backgrounds, we synthesized all possible genotypic intermediates in the line of descent connecting the wildtype bar-headed goose genotype with the most recent common ancestor of bar-headed goose and its lowland relatives. Site-directed mutagenesis experiments revealed one major-effect mutation that significantly increased Hb-O2 affinity on all possible genetic backgrounds. Two other mutations exhibited smaller average effect sizes and less additivity across backgrounds. One of the latter mutations produced a concomitant increase in the autoxidation rate, a deleterious side-effect that was fully compensated by a second-site mutation at a spatially proximal residue. The experiments revealed three key insights: (i) subtle, localized structural changes can produce large functional effects; (ii) relative effect sizes of function-altering mutations may depend on the sequential order in which they occur; and (iii) compensation of deleterious pleiotropic effects may play an important role in the adaptive evolution of protein function.

Comparative analyses of the gut microbiota among three different wild geese species in the genus Anser.

In this study, we characterized for the first time the gut microbiota of Greylag geese (Anser anser) using high-throughput 16S rRNA gene sequencing technology. The results showed that the phyla Firmicutes (78.55%), Fusobacteria (9.38%), Proteobacteria (7.55%), Bacteroidetes (1.82%), Cyanobacteria (1.44%), and Actinobacteria (0.61%) dominated the gut microbial communities in the Greylag geese. Then, the variations of gut microbial community structures and functions among the three geese species, Greylag geese, Bar-headed geese (Anser indicus), and Swan geese (Anser cygnoides), were explored. The greatest gut microbial diversity was found in Bar-headed geese group, while other two groups had the least. The dominant bacterial phyla across all samples were Firmicutes and Proteobacteria, but several characteristic bacterial phyla and genera associated with each group were also detected. At all taxonomic levels, the microbial community structure of Swan geese was different from those of Greylag geese and Bar-headed geese, whereas the latter two groups were less different. Functional KEGG categories and pathways associated with carbohydrate metabolism, energy metabolism, and amino acid metabolism were differentially expressed among different geese species. Taken together, this study could provide valuable information to the vast, and yet little explored, research field of wild birds gut microbiome.

A history of hybrids? Genomic patterns of introgression in the True Geese.

BACKGROUND: The impacts of hybridization on the process of speciation are manifold, leading to distinct patterns across the genome. Genetic differentiation accumulates in certain genomic regions, while divergence is hampered in other regions by homogenizing gene flow, resulting in a heterogeneous genomic landscape. A consequence of this heterogeneity is that genomes are mosaics of different gene histories that can be compared to unravel complex speciation and hybridization events. However, incomplete lineage sorting (often the outcome of rapid speciation) can result in similar patterns. New statistical techniques, such as the D-statistic and hybridization networks, can be applied to disentangle the contributions of hybridization and incomplete lineage sorting. We unravel patterns of hybridization and incomplete lineage sorting during and after the diversification of the True Geese (family Anatidae, tribe Anserini, genera Anser and Branta) using an exon-based hybridization network approach and taking advantage of discordant gene tree histories by re-sequencing all taxa of this clade. In addition, we determine the timing of introgression and reconstruct historical effective population sizes for all goose species to infer which demographic or biogeographic factors might explain the observed patterns of introgression. RESULTS: We find indications for ancient interspecific gene flow during the diversification of the True Geese and were able to pinpoint several putative hybridization events. Specifically, in the genus Branta, both the ancestor of the White-cheeked Geese (Hawaiian Goose, Canada Goose, Cackling Goose and Barnacle Goose) and the ancestor of the Brent Goose hybridized with Red-breasted Goose. One hybridization network suggests a hybrid origin for the Red-breasted Goose, but this scenario seems unlikely and it not supported by the D-statistic analysis. The complex, highly reticulated evolutionary history of the genus Anser hampered the estimation of ancient hybridization events by means of hybridization networks. The reconstruction of historical effective population sizes shows that most species showed a steady increase during the Pliocene and Pleistocene. These large effective population sizes might have facilitated contact between diverging goose species, resulting in the establishment of hybrid zones and consequent gene flow. CONCLUSIONS: Our analyses suggest that the evolutionary history of the True Geese is influenced by introgressive hybridization. The approach that we have used, based on genome-wide phylogenetic incongruence and network analyses, will be a useful procedure to reconstruct the complex evolutionary histories of many naturally hybridizing species groups.

Are declining populations of wild geese in China 'prisoners' of their natural habitats?

While wild goose populations wintering in North America and Europe are mostly flourishing by exploiting farmland, those in China (which seem confined to natural wetlands) are generally declining. Telemetry devices were attached to 67 wintering wild geese of five different species at three important wetlands in the Yangtze River Floodplain (YRF), China to determine habitat use. 50 individuals of three declining species were almost entirely diurnally confined to natural wetlands; 17 individuals from two species showing stable trends used wetlands 83% and 90% of the time, otherwise resorting to farmland. These results confirm earlier studies linking declines among Chinese wintering geese to natural habitat loss and degradation affecting food supply. These results also contribute to explaining the poor conservation status of Chinese wintering geese compared to the same and other goose species wintering in adjacent Korea and Japan, western Europe and North America, which feed almost entirely on agricultural land, liberating them from winter population limitation.

Transiently expressed pattern during myogenesis and candidate miRNAs of Tmem8C in goose.

Transmembrane protein 8C (Tmem8C) is a muscle-specific membrane protein that controls myoblast fusion, which is essential for the formation of multinucleated muscle fibres. As most of the birds can fly, they have enormous requirement for the muscle, but there are only a few studies of Tmem8C in birds. In this study, we obtained the coding sequence (CDS) of Tmem8C in goose, predicted miRNAs that can act on the 3'UTR, analysed expression profiles of this gene in breast and leg muscles (BM and LM) during the embryonic period and neonatal stages, and identified miRNAs that might affect the targeted gene. The results revealed a high homology between Tmem8C in goose and other animals (indicated by sequence comparisons and phylogenetic trees), some conservative characteristics (e.g., six transmembrane domains and two E-boxes in the 5'UTR might be the potential binding sites of muscle regulatory factors (MRFs)), and the dN/dS ratio indicated purifying selection acting on this gene, facilitating conservatism in vertebrates. Q-PCR indicated Tmem8C had a peak expression pattern, reaching its highest expression levels in stage E15 in LM and E19 in BM, and then dropping transiently in E23 (P < 0.05). We examined 13 candidate miRNAs, and negative relationships were detected both in BM and LM (mir-125b-5p, mir-15a, mir-16-1 and mir-n23). Notably, mir-16-1 significantly decreased luciferase activity in dual luciferase reporter gene (LRG) assay, suggesting that it can be identified as potential factors affecting Tmem8C. This study investigated Tmem8C in water bird for the first time, and provided useful information about this gene and its candidate miRNAs in goose.

Identification of 2'-5'-Oligoadenylate Synthetase-Like Gene in Goose: Gene Structure, Expression Patterns, and Antiviral Activity Against Newcastle Disease Virus.

2'-5'-oligoadenylate synthetase-like (OASL) is a kind of antiviral protein induced by interferons (IFNs), which plays an important role in the IFNs-mediated antiviral signaling pathway. In this study, we cloned and identified OASL in the Chinese goose for the first time. Goose 2'-5'-oligoadenylate synthetase-like (goOASL), including an ORF of 1527bp, encoding a protein of 508 amino acids. GoOASL protein contains 3 conserved motifs: nucleotidyltransferase (NTase) domain, 2'-5'-oligoadenylate synthetase (OAS) domain, and 2 ubiquitin-like (UBL) repeats. The tissue distribution profile of goOASL in 2-week-old gosling and adult goose were identified by Real-Time quantitative PCR, which revealed that the highest level of goOASL mRNA transcription was detected in the blood of adult goose and gosling. The mRNA transcription level of goOASL was upregulated in all tested tissues of duck Tembusu virus (DTMUV)-infected 3-day-old goslings, compared with control groups. Furthermore, using the stimulus Poly(I: C), ODN2006, R848, and lipopolysaccharide (LPS) as well as the viral pathogens DTMUV, H9N2 avian influenza virus (AIV), and gosling plague virus (GPV) to treat goose peripheral blood mononuclear cells (PBMCs) for 6 h, goOASL transcripts level was significantly upregulated in all treated groups. To further investigate the antiviral activity of goOASL, pcDNA3.1(+)-goOASL-His plasmid was constructed, and goOASL was expressed by the goose embryo fibroblast cells (GEFs) transfected with pcDNA3.1(+)-goOASL-His. Our research data suggested that Newcastle disease virus (NDV) replication (viral copies and viral titer) in GEFs was significantly reduced by the overexpression of goOASL protein. These data were meaningful for the antiviral immunity research of goose and shed light on the future prevention of NDV in fowl.

A tree of geese: A phylogenomic perspective on the evolutionary history of True Geese.

Phylogenetic incongruence can be caused by analytical shortcomings or can be the result of biological processes, such as hybridization, incomplete lineage sorting and gene duplication. Differentiation between these causes of incongruence is essential to unravel complex speciation and diversification events. The phylogeny of the True Geese (tribe Anserini, Anatidae, Anseriformes) was, until now, contentious, i.e., the phylogenetic relationships and the timing of divergence between the different goose species could not be fully resolved. We sequenced nineteen goose genomes (representing seventeen species of which three subspecies of the Brent Goose, Branta bernicla) and used an exon-based phylogenomic approach (41,736 exons, representing 5887 genes) to unravel the evolutionary history of this bird group. We thereby provide general guidance on the combination of whole genome evolutionary analyses and analytical tools for such cases where previous attempts to resolve the phylogenetic history of several taxa could not be unravelled. Identical topologies were obtained using either a concatenation (based upon an alignment of 6,630,626 base pairs) or a coalescent-based consensus method. Two major lineages, corresponding to the genera Anser and Branta, were strongly supported. Within the Branta lineage, the White-cheeked Geese form a well-supported sub-lineage that is sister to the Red-breasted Goose (Branta ruficollis). In addition, two main clades of Anser species could be identified, the White Geese and the Grey Geese. The results from the consensus method suggest that the diversification of the genus Anser is heavily influenced by rapid speciation and by hybridization, which may explain the failure of previous studies to resolve the phylogenetic relationships within this genus. The majority of speciation events took place in the late Pliocene and early Pleistocene (between 4 and 2millionyears ago), conceivably driven by a global cooling trend that led to the establishment of a circumpolar tundra belt and the emergence of temperate grasslands. Our approach will be a fruitful strategy for resolving many other complex evolutionary histories at the level of genera, species, and subspecies.

Molecular cloning of the SMAD4 gene and its mRNA expression analysis in ovarian follicles of the Yangzhou goose (Anser cygnoides).

Mothers against decapentaplegic homolog 4 (SMAD4) is an important protein in animal reproduction. It plays pivotal roles in cellular pathways, including apoptosis. The expression profile of the SMAD4 gene in goose ovarian follicles has not been reported. In this study, the SMAD4 coding sequence was cloned from the Yangzhou goose. A phylogenetic analysis was performed and mRNA expression was examined in various tissues using quantitative real-time PCR. An alternative splice form of SMAD4, SMAD4-b having 1656 bp, was identified. SMAD4-a mRNA was widely expressed in various healthy tissues, whereas SMAD4-b was very weakly expressed. SMAD4 mRNA in the ovary and oviduct was significantly higher than that in the pituitary and hypothalamus. SMAD4 mRNA expression analysis in hierarchical follicles showed that the level of SMAD4 mRNA was higher in large white follicles and post-ovulatory follicles than in the other follicles. The results indicate that SMAD4 might be involved in the recruitment of hierarchical follicles.

Complete mitochondrial genome of the Swan Goose (Anser cygnoides L.) and its phylogenetic analysis.

In this study, we undertook the first complete the Swan Goose (Anser cygnoides L.) mitochondrial genome. The total length of the mitogenome is 16,739 bp. It contains the typical structure, including 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes, and one D-loop region. The overall composition of the mitogenome is A (29.2%), G (15.8%), C (32.8%), and T (22.2%). Besides, the base preference of AT was not determined. Twenty-two kinds of tRNA were all typically cloverleaf structures. According to the phylogenetic analysis, A. cygnoides L. has a closer relationship with Anser anser.

Phylogenetic studies of four Anser cygnoides (Anserini: Anserinae) in Hunan province of China based on complete mitochondrial DNA sequences.

In this study, we cloned and sequenced the complete mitochondrial DNAs of Chinese goose, Anser cygnoides populations from three different areas of Hunan province in China. The Anser cygnoides breed Wugangtong white goose (WGTW) sample and Wugangtong grey goose sample (WGTG) were taken from the Wugang county of Shaoyang city, the Anser cygnoides breed Xupu goose (XP) sample was taken from the Xupu county of Huaihua city, and the Anser cygnoides breed Yanling white goose (YLW) sample was taken from the Yanling county of Zhuzhou city. The organization of the four Anser cygnoides breeds mitochondrial genomes was similar. Phylogenetic analyses using N-J computational algorithms showed that the analyzed species are divided into four major clades: Anatinae, Anserinae, Dendrocygninae and Anseranatidae. It was noted that Wugangtong white goose, Yanling white goose and Xupu goose have highly similar phylogenetic relationship.

Molecular cloning and mRNA expression analysis of ornithine decarboxylase antizyme 2 in ovarian follicles of the Sichuan white goose (Anser cygnoides).

The ornithine decarboxylase antizyme 2 (OAZ2) gene is a member of the antizyme gene family. Antizymes play pivotal roles in various cellular pathways, including polyamine anabolism and apoptosis. The molecular structure and expression profile of the OAZ2 in goose ovarian follicles have not been reported. In this study, the OAZ2 cDNA sequence of the Sichuan white goose was cloned (Anser cygnoides), and phylogenetic and structural analyses of the OAZ2 were performed. The expression profiling of OAZ2 mRNA in goose ovarian follicles was examined using quantitative real-time PCR. The sequence analysis showed that the 756 bp OAZ2 sequence contained two overlapping open reading frames (ORF). ORF1 was 99 bp in length, and encoded a 32 aa polypeptide. ORF2 was 477 bp in length, and encoded a 158 aa polypeptide. The frameshift site that initiates the translation of ORF2 was located at nucleotide position 97 in the OAZ2. The analysis of OAZ2 mRNA expression in hierarchical follicles showed that the level of OAZ2 mRNA was higher in the SWF and F2 follicular stages than that in the ovarian stroma (P<0.05). The lowest level of OAZ2 expression was detected in the ovarian stroma. These results suggest that the highly conserved frameshift region plays an important role in sustaining the function of OAZs. Furthermore, the significantly higher level of OAZ2 mRNA in the SWF stage indicates that OAZ2 may be involved in recruiting hierarchical follicles. Our results also suggest that OAZ2 may augment the effects of OAZ1 in follicle development.

Molecular characterisation and expression profiling of the ENO1 gene in the ovarian follicle of the Sichuan white goose.

The ENO1 gene encodes a multifunctional enzyme that has been identified as a key component of the glycolytic pathway. Our previous studies demonstrated that ENO1 gene expression was higher in the ovaries of laying geese compared with prelaying geese. However, the molecular characterisation and expression profiling of the ENO1 gene in geese tissues and ovarian follicles remain to be determined. In this study, ENO1 cDNA (1,445 bp long) of the Sichuan white goose was cloned and characterised. The ORF of ENO1 cDNA is 1,305 bp in length and encodes a 434 amino acid protein with a molecular weight of 47.27 kDa. ENO1 expression in all of the examined tissues was the highest in spleen and the lowest in breast muscle. High expression of ENO1 appeared in the kidney, liver, adrenal gland, and retina. With increasing follicle growth, ENO1 gene expression began to decrease from the small white follicle to F5, which was followed by a sharp increase in expression in F4 and then a gradual decrease in expression from F3 to F1. Furthermore, in the postovulatory follicles (POF), the levels of ENO1 gene expression decreased gradually from POF1 to POF4. In conclusion, the ENO1 transcript was widely distributed in various tissues of the Sichuan white goose, but ENO1 expression was tissue-specific. Furthermore, the results of the ENO1 expression profiling of ovarian follicles suggest that ENO1 may play an important dual role in the progress of follicular development, where ENO1 acts as a glycolytic enzyme and also mediates apoptosis.

Molecular characterization and differential expression of multiple goose dopamine D2 receptors.

Dopamine D2 receptor (DRD2) gene, a member of the dopamine receptors gene family, has been studied as a candidate gene for broodiness due to its special effects on avian prolactin secretion. Here, the genomic DNA and cDNA sequences of goose (Anser cygnoides) DRD2 gene were cloned and characterized for the first time. The goose DRD2 cDNA is 1353bp in length and encodes a protein of 450 amino acids. The length of goose DRD2 genomic DNA is 8350bp, including seven exons and six introns. We identified four goose DRD2 variants, which were generated due to alternative splicing. Bioinformatics analysis indicates that all the deduced DRD2 amino acid sequences contain seven putative transmembrane domains and four potential N-glycosylation sites. A phylogenetic tree based on amino acid sequences displays that the goose DRD2 protein is closely related to those of avian species. Semi-quantitative RT-PCR analysis demonstrates that the DRD2-1, DRD2-2 and DRD2-4 transcripts are differentially expressed in the pituitary, ovary, hypothalamus, as well as in the kidney, whereas the DRD2-3 transcript is widely expressed in all the examined tissues at different levels. Meanwhile, 54 single nucleotide polymorphisms (SNPs) and 4 insert-deletion (indel) variations were identified in the coding region and partial intron region of the goose DRD2 gene. Those findings will help us gain insight into the functions of the DRD2 gene in geese.