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【目的】构建产蛋前后各期伊犁鹅卵巢转录组文库,结合生物信息学分析揭示不同产蛋期伊犁鹅卵巢组织差异表达基因,并鉴定出影响鹅卵巢发育的关键基因,为伊犁鹅的繁殖调控提供理论参考。【方法】选择处于开产期（KL组）、产蛋期（CL组）及休产期（XL组）的伊犁鹅各4只,屠宰后取其卵巢组织,用于构建卵巢转录组文库。通过新基因挖掘、基因差异表达、基因注释以及蛋白互作网络分析筛选出与鹅卵泡发育相关的候选基因;随机挑选8个差异表达基因,应用实时荧光定量PCR验证其表达情况。【结果】伊犁鹅卵巢组织学结果表明,在开产期时,伊犁鹅卵巢表面有大量初级卵泡,而产蛋期卵巢则显示出卵泡的层级性,在休产期卵巢中可观察到卵泡出现向内凹陷、闭锁的现象。通过转录组测序（RNA-Seq）,从构建的12个伊犁鹅卵巢cDNA文库中获得有效读数57 811 186~85 328 377条,Q30值均>93.38%,每个样品所产的测序读数于鹅参考基因组上的比对率在82.79%~89.24%。在卵巢中注释的新基因共有1 112个,单核苷酸多态性（SNP）位点总数为1 642 273~2 425 069个;SNP和插入缺失（InDel）均主要注释于内含子区域;各时期的可变剪切类型主要集中为最后1个外显子可变剪切（TSS）及第1个外显子可变剪切（TTS）。在KL vs CL、XL vs CL及XL vs KL组中分别有337、1 136和525个差异表达基因,共有差异表达基因为α2A肾上腺素能受体（ADRA2A）、表皮蛋白（CP）、非转移性黑色素瘤糖蛋白B （GPNMB）及α-1-抗胰蛋白酶样（LOC106033756）。GO功能富集分析发现,差异表达基因主要富集在肽基酪氨酸磷酸化的正调控、细胞黏附及质膜外侧等过程。KEGG通路富集分析发现,差异表达基因主要显著富集于神经活性配体-受体相互作用、ECM-受体相互作用及类固醇生物合成等。结合蛋白互作网络分析,筛选到与鹅卵巢发育相关的潜在调控因子Bruton’s酪氨酸激酶（BTK）、血小板源性生长因子受体α（PDGFRA）、整合素β3（ITGB3）等。实时荧光定量PCR结果显示,RNA-Seq结果准确可靠。【结论】本研究揭示了产蛋前后各期伊犁鹅卵巢组织中的基因表达差异,筛选到影响伊犁鹅卵巢发育的神经活性配体-受体相互作用、ECM-受体相互作用、类固醇生物合成等重要通路与BTK、PDGFRA和ITGB3等关键候选基因,为了解鹅卵巢组织调控产蛋性能的分子机制提供了理论支撑。 相似文献
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Comparative proteomic analysis of hypothalamus tissue from Huoyan geese between pre‐laying period and laying period using an iTRAQ‐based approach 
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下载免费PDF全文 Zhongzan Cao Ruiming Fan Bo Meng Zhe Xing Mei Liu Ming Gao Xinhong Luan 《Animal Science Journal》2018,89(7):946-955
The hypothalamus plays a central role in controlling poultry endocrine and reproductive activities. So far there is limited information focused on the proteome profiles of the hypothalamus from geese during different stages of the egg‐laying cycle. In order to identify proteins regulating the egg‐laying process of Huoyan geese, we investigated the proteome profiles of the hypothalamus from Huoyan geese during the laying period and pre‐laying period by applying an isobaric tags for relative and absolute quantitation (iTRAQ)‐based proteomic technology. A total number of 3,337 were identified and quantified, of which 18 were significantly up‐regulated and 16 were significantly down‐regulated. These differentially expressed proteins were subjected to bioinformatics analyses based on the Gene Ontology annotation and Kyoto Encyclopedia of Genes and Genomes pathway. Some of these were revealed to be involved in hormone and neurotransmitter secretion, exocytosis, calcium ion transport and synaptic transmission. Subsequently, excitatory amino acid transporter 2, complexin‐1 and inositol 1,4,5‐trisphosphate receptor, type 3 were confirmed at the messenger RNA level using quantitative real‐time RT‐PCR. Then, the abundance change of these proteins was verified further using Western blotting analysis. These data may aid in elucidating the molecular mechanism of higher laying performance in Huoyan geese. 相似文献
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采用实时定量PCR技术定量检测产蛋前期和产蛋期籽鹅卵巢组织中铁蛋白重链基因(FTH)和8个新ESTs(ODEUG01~ODEUG08)的mRNA表达水平。结果表明:产蛋期籽鹅卵巢组织中FTH和8个新ESTs的相对表达量分别比产蛋前期卵巢组织高13.25、7.68、11.82、14.23、8.25、15.14、9.78、6.48、14.21倍。本研究进一步证实,FTH和8个新ESTs在产蛋期籽鹅卵巢组织中高效表达,FTH和8个新ESTs可能参与籽鹅卵巢功能的调节,并影响籽鹅的产蛋性能。 相似文献
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Zhixiu Wang Laidi Wang Yang Zhang Ying Yao Wenming Zhao Qi Xu Guohong Chen 《Journal of animal physiology and animal nutrition》2021,105(5):938-945
Zhedong white goose (Anser cygnoides domesticus) is a native Chinese breed with strong broodiness and low egg production, which is related to the physiology of reproduction. However, thus far, the physiology of goose reproduction has not been well elucidated. In the present study, the ovarian morphology and reproductive hormones of Zhedong white geese were investigated during the reproductive cycle (the laying and brooding periods). The results showed that the surface of the ovary was atrophied and follicular atresia appeared to some extent in the brooding period compared with the laying period. The concentrations of follicle-stimulating hormone, progesterone and luteinizing hormone were significantly higher than those in the brooding period (p < 0.05). In contrast, the concentrations of prolactin (PRL) and anti-Müllerian hormone (AMH) in the laying period were significantly lower than those in the brooding period (p < 0.05). In addition, the mRNA expression levels of PRL, AMH, dopamine-β-hydroxylase (DβH) and cytochrome P450 side-chain cleavage enzyme (P450scc) were detected in the hypothalamus, pituitary and ovaries by using real-time polymerase chain reaction. The results showed that AMH mRNA was expressed specifically in ovary tissue. The expression levels of DβH and PRL in the brooding period was significantly higher than those in the laying period in the three tissues, especially in the early and middle stages of the brooding period. Moreover, AMH mRNA expression in the ovaries presented the same trend. In addition, P450scc mRNA was highly expressed in both the ovary and pituitary in the laying period. These results revealed the remarkable features of ovarian morphology and characterized the hormonal pattern and expression profile during the reproductive cycle, all of which contribute to understanding the differences in reproductive physiology between the laying and brooding periods in Zhedong white geese. 相似文献
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C Yang J Ye Y Liu J Ding H Liu X Gao X Li Y Zhang J Zhou X Zhang W Huang F Fang Y Ling 《Reproduction in domestic animals》2018,53(3):793-800
Puberty is initiated by increased pulsatile gonadotropin‐releasing hormone (GnRH) release from the hypothalamus. Epigenetic repression is thought to play a crucial role in the initiation of puberty, although the existence of analogous changes in methylation patterns across species is unclear. We analysed mRNA expression of DNA methyltransferases (DNMTs) and methyl‐binding proteins (MBPs) in goats and rats by quantitative real‐time PCR (qRT‐PCR). DNA methylation profiles of hypothalamic were determined at the pre‐pubertal and pubertal stages by bisulphite sequencing. In this study, expression of DNMTs and MBPs mRNA showed different patterns in goats and rats. Global methylation variation was low in goats and rats, and the profile remained stable during puberty. Gene ontology (GO) and Kyoto Encyclopedia of Gene and Genomes (KEGG) pathway analysis revealed the involvement of 62 pathways in puberty in goats and rats including reproduction, type I diabetes mellitus and GnRH signalling pathways and found that Edn3, PTPRN2 and GRID1 showed different methylation patterns during puberty in goats and rats and similar variation patterns for Edn3 and PTPRN2 were showed. These indicated that Edn3 and PTPRN2 would play a role in the timing of puberty. This study provides evidence of the epigenetic control of puberty. 相似文献
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Xiaodong Zhang Liang Zhang Jinnan Shang Qiangqiang Tao Mi Tian Yingchun Ma Yiliang Xu Yueyun Ding Rong Zhou Kui Li Zongjun Yin 《Reproduction in domestic animals》2019,54(7):1018-1025
The aim of this study was to explore the expression difference of miRNAs and mRNAs between the follicular phase (FP) and luteal phase (LP) in porcine ovaries and provide a theoretical basis for the research on mammalian reproductive regulation. RNA‐Seq and miRNA‐Seq were used to identify differentially expressed genes (DEGs) and miRNAs (DEMs) between the FP and LP in ovaries of six sows (3‐year‐old Yorkshire pigs with similar weights and same parities). Bioinformatic analysis was used to screen potential genes and miRNAs related to porcine ovarian function. Real‐time qualitative PCR was used to validate the sequencing results. RNA‐Seq results showed that 3,078 genes were up‐regulated, and 1,444 genes were down‐regulated in the LP compared with the FP, and DEGs were significantly enriched in 242 Gene Ontology (GO) terms and 33 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. miRNA‐Seq identified 112 DEMs, of which 25 were up‐regulated and 87 were down‐regulated in the LP compared with the FP. We obtained 186 intersection genes (IGs) between the 4,522 DEGs and 2,444 target genes predicted from the 112 DEMs. After constructing a miRNA‐gene‐pathway network, we identified key miRNAs and genes including miR‐17‐3p, miR‐214, miR‐221‐5p, miR‐125b, FGF1, YWHAG, YWHAZ, FDFT1 and DHCR24, which are enriched in Hippo and PI3K‐Akt signalling pathways, and various metabolic pathways. These results indicate that these key genes and miRNAs may play important roles in the developmental transition from FP to LP in porcine ovaries and represent candidate targets for further study. 相似文献
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Naseer A. Baba Manjit Panigrahi Ankita D. Verma Abdul Sadam Sourabh Sulabh Supriya Chhotaray Subhashree Parida Narayanan Krishnaswamy Bharat Bhushan 《Reproduction in domestic animals》2019,54(1):100-107
Progesterone (P4) plays a key role in the establishment and maintenance of pregnancy in most mammals. Unravelling the expression of progesterone‐regulated genes can expand the understanding of the embryonic mortality. Accordingly, we studied the relative mRNA expression of the P4‐regulated genes in the buffalo. Uteri were collected from the abattoir and categorized into nonpregnant late luteal phase, stage I (28–38th days of gestation) and stage II (48–56th days of gestation) of pregnancy (n = 6/group). After extraction of total RNA from the endometrial tissues, we carried out qRT‐PCR for determining the relative mRNA expression of the P4‐regulated genes using nonpregnant late luteal phase as calibrator group. The expression of LGALS3BP (essential for maternal recognition of pregnancy) gene was found to be significantly upregulated (p < 0.05), while MUC1 (important for embryo attachment) gene was downregulated in stage I and II of pregnancy. We observed no significant change in the expression of LGALS1, LGALS9 and CTSL genes. The SLC5A11 and SLC2A1 genes (involved in the transport of glucose to endometrium) in early pregnancy were upregulated in the pregnancy stage I (p < 0.05) relative to nonpregnant late luteal phase. The CST3 gene was significantly upregulated in pregnancy stage II (p < 0.01). These results provide molecular insights into the specific pathways involved in foeto‐maternal communication during early pregnancy in buffaloes. 相似文献
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Haiying CHI Masahiro SATO Mitsutoshi YOSHIDA Kazuchika MIYOSHI 《Animal Science Journal》2012,83(1):88-93
α‐1,3‐Galactosyltransferase (α‐GalT), an enzyme creating Galα1‐3Gal (α‐Gal) epitope on the cell surface in some mammalian species such as pigs, is known to be a key factor that causes hyperacute rejection upon transplantation from pigs to humans. To establish the RNA interference‐based suppression of endogenous α‐GalT messenger RNA (mRNA) synthesis in porcine preimplantation embryos, we determined the suitable embryonic stage at which stage such approach is possible by using the semi‐quantitative RT‐PCR (qRT‐PCR) and the cytochemical method using a fluorescence‐labeled Bandeiraea simplicifolia Isolectin B4 (BS‐I‐B4). Staining with BS‐I‐B4 demonstrated that α‐Gal epitope expression was first recognized at the 8‐cell stage, and increased up to the hatched blastocyst stage. Single embryo‐based qRT‐PCR also confirmed this pattern. These results indicate that creation of α‐Gal epitope is proceeded by de novo synthesis of α‐GalT mRNA in porcine preimplantation embryos with peaking at the blastocyst stage. 相似文献
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Here, we describe the establishment of mutant‐specific polymerase chain reaction (PCR) for detection of a c‐KIT c.1430G>T mutation in feline mast cell tumours. Several mutations in feline c‐KIT have been identified, with the c.1430G>T mutation accounting for a significant portion of feline mast cell tumour mutations. The c.1430G>T mutation in c‐KIT exon 9 was detected in 15.7% (11 of 70) of samples by mutant‐specific PCR but in only 7.1% (5 of 70) by PCR–restriction fragment length polymorphism (RFLP) in the genomic DNA isolated from 70 formalin‐fixed paraffin‐embedded sections or cells collected by fine needle aspiration. Mutant‐specific PCR showed remarkably higher detection rate than did PCR–RFLP. DNA sequence analysis did not always yield identical results to those of mutant‐specific PCR, suggesting heterogeneity of tumour cells. Mutant‐specific PCR is a valid and efficient screening tool for detection of the c‐KIT c.1430G>T point mutation in feline mast cell tumours compared with PCR–RFLP and sequencing analysis. 相似文献
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Gonadotropin-releasing hormone (GnRH), a neuropeptide, plays a vital role in the hypothalamus–pituitary–gonadal (HPG) axis. In vertebrates, GnRH is crucial for the onset of sexual development and the entire reproductive process. The purpose of this study was to identify genetic factors associated with egg-laying traits of Muscovy ducks.
The full-length cDNA (474 bp) of Muscovy duck GnRH was obtained and characterised. It encodes 92 amino acids containing a 1-amino acid signal peptide cleavage site. Phylogenetic analysis revealed that Muscovy duck GnRH has a close relationship with Anas platyrhynchos GnRH.
GnRH showed significantly different expression profiles between 4 developmental periods in the hypothalamus, pituitary, and ovary. The expression of GnRH in the laying period (36 weeks) was higher than at other periods in the three tissues. GnRH was widely expressed in 12 examined tissues of nesting and laying Muscovy ducks. In the hypothalamus, pituitary and gonads, the expression of GnRH was higher than in other tissues.
In laying Muscovy ducks, the expression of GnRH in the hypothalamus, pituitary, ovary, muscular stomach, pancreas, heart, duodenum and spleen was significantly higher than in nesting dusks. Differences were detected in the liver and glandular stomach between laying ducks and nesting ducks. Differences between the kidney and lung were not significant.
In the pituitary, the GnRH and GnRH receptor (GnRHR) genes shared the same expression profiles during 4 time points. Both genes had the highest expression at 36 weeks of age.
A mutation (g.206G > A) in the 5?-flanking region was associated with egg-laying performance. Individuals with genotype GG had better egg-laying performance than the individuals with genotype AA. GnRH may be used as a marker gene for laying performance in the Muscovy duck.
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