Supranutritional dietary selenium depressed expression of selenoprotein genes in three immune organs of broilers

The objective of this study was to investigate the effects of supranutritional dietary selenium (Se) on selenoproteins expression in three immune organs of chickens. A total of 160 1‐day‐old male Cobb broilers were randomly divided into two groups and fed a Se‐deficient corn‐soybean basal diet supplemented with 0.3 (adequate) and 3.0 (excess) mg/kg Se for 42 days. Immune organs were collected, and effects of supranutritional Se on messenger RNA abundance of 23 selenoprotein genes and eight inflammation‐related genes were compared at day 42. Also enzyme activities were measured at days 14, 28 and 42. The results showed supranutritional dietary Se depressed growth performance of chicken and down‐regulated nine and three selenoprotein genes in thymus and spleen, respectively, and only Sepp1 was up‐regulated in the bursa of Fabricius. Also three, three and seven inflammation‐related genes were up‐regulated in three organs, respectively. Supranutritional Se elevated (P < 0.05) activities of superoxidase dismutase, total antioxidant capacity and glutathione peroxidase, mainly in early stages. In summary, supranutritional Se resulted in down‐regulation of selenoprotein genes and up‐regulation of inflammation‐related genes in three immune organs of chicken, which indicated potential roles of those selenoprotein genes in immune organs of the chicken.     

Comparative proteomic analysis of hypothalamus tissue from Huoyan geese between pre‐laying period and laying period using an iTRAQ‐based approach

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.     

Th1/Th2型免疫反应相关基因在巨噬细胞RAW 264.7应对细粒棘球绦虫原头蚴刺激时的表达谱研究

本研究旨在解析巨噬细胞RAW264.7在应对细粒棘球绦虫原头蚴刺激时其Th1、Th2型免疫反应相关基因的差异表达规律,为进一步揭示巨噬细胞抗细粒棘球绦虫原头蚴免疫调控机制奠定理论基础.将细粒棘球绦虫原头蚴和巨噬细胞RAW264.7共培养6、24、72 h,收集RAW264.7细胞,提取总RNA,构建cDNA文库,利用R...     

冷应激后民猪背脂中差异lncRNA的筛选与分析

旨在分析冷应激对猪脂肪组织内lncRNA表达的影响,探讨冷应激下lncRNA与靶基因的改变对脂肪代谢及机体的影响。本研究以6头6月龄雌性民猪为试验材料,随机分为2组,一组置于正常舍内饲养,温度控制在（18±2）℃,设为对照组;另一组置于室外半敞篷舍内饲养,温度控制在（-18±5）℃,设为试验组。处理24 h后,屠宰,取背部皮下脂肪为试验材料,通过转录组测序筛选出受低温诱导的lncRNA及其靶基因,并对差异基因开展生物学功能分析,构建两者间的调控网络。结果发现,166个差异表达lncRNAs和269个基因受到冷应激诱导,其中仅有2个为已知的lncRNAs。差异基因中有多个与机体免疫相关,如干扰素刺激基因、DHX58、PTX3和OASL等,与呼吸、氧化应激和血液循环相关的基因有KCNK3、CCDC38、GAS2L2、KNDC1、LIPG和C1QL3等,与脂类代谢相关的基因有ANGPTL4、KLF11、NUPR1和HERC5,它们均发生了显著上调。而与神经系统相关的基因,如SLIT1、MAP6、TREM2和IRFN2等则发生了显著下调。差异基因参与的生物学过程主要包括防御反应、对生物刺激的反应以及病毒的反应等。通过相关系数分析发现,144个lncRNA与208个靶基因共表达,并以反式调控为主,两者间存在着复杂的调控关系,所筛选出的靶基因显著富集到真核生物的核糖体生物发生和基底细胞癌通路。据此推测,冷应激下机体的先天免疫系统、呼吸、氧化应激、血液循环和神经系统的敏感性等生理过程均受到影响,脂类代谢发生了改变。     

Long‐term dietary supplementation of organic selenium modulates gene expression profiles in leukocytes of adult pigs

Seventy‐two pigs at 34.4 kg body weight (BW) were allotted to two treatments with six replicates/treatment and six pigs/pen: the CON (negative control, no added selenium (Se)) and the OS (0.36 mg/kg added selenium from selenium‐enriched yeast). Pigs were fed until 130 kg BW. The CON diet contained 0.18 mg/kg indigenous Se whereas the OS diet contained 0.54 mg/kg Se. Blood samples were collected at 130 kg BW and further processed for microarray analysis, prepared with 885 genes related to immune function of pigs. Among those, 28 genes related to improved immune status and innate immunity were up‐regulated (P < 0.05) in leukocytes from Se‐fed pigs and those include major histocompatibility class I (> 1.66), arginase I (> 1.27), integrin beta‐1‐subunit (> 1.20), toll like receptor 2 (> 1.12) and double‐stranded RNA‐dependent protein kinase. However, 24 genes including tissue factor (< 4.70), serum amyloid A‐2 protein (< 3.11) and p27Kip1 (< 1.42) were down‐regulated (P < 0.05) in leukocytes from Se‐fed pigs. Expression of four selected genes was validated using quantitative PCR (qPCR) showing significant correlation between mircroarray analysis and qPCR analysis. This study indicates that a long‐ term dietary supplementation (0.3%) of organic Se improves the expression of genes that are related to enhanced immunity of pigs.     

伊犁鹅产蛋前后各期卵巢转录谱的构建及卵泡发育相关基因的分析

【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。     

非洲猪瘟病毒感染后猪外周血淋巴细胞Toll样受体信号通路的lncRNA-mRNA分析

【目的】丰富非洲猪瘟病毒（African swine fever virus,ASFV）感染后猪外周血淋巴细胞长链非编码RNA（long non-coding RNA,lncRNA）表达谱,并进一步挖掘影响Toll样受体信号通路的调控网络。【方法】试验动物感染ASFV,于第7天采集外周血并分离得到外周血淋巴细胞,运用Illumina高通量组学测序对外周血淋巴细胞中lncRNA进行测序,原始数据经处理后筛选获得差异表达的lncRNA,并进行靶基因预测,利用生物信息学方法对靶基因进行GO功能和KEGG信号通路富集分析,初步绘制与Toll样受体信号通路相关的lncRNA-mRNA调控网络,并对lncRNA-ENSSSCG00000041959在内的4个lncRNAs进行实时荧光定量RT-PCR验证。【结果】共筛选到73个差异表达的lncRNAs,其中上调表达lncRNAs 38个,下调表达lncRNAs 35个。GO功能分析结果显示,靶基因显著富集在调节免疫系统过程、防御反应、生物刺激、病毒反应和先天性免疫;KEGG信号通路富集分析显示,大部分靶基因与细胞循环、疾病及免疫应答有关,其中免疫相关信号通路有Toll样受体信号通路、TNF信号通路、产生IgA的肠道免疫网络等。进一步挖掘出lncRNA-ENSSSCG00000041959-RIPK1和lncRNA-ENSSSCG00000041959-IRAK1可能是影响Toll样受体信号通路的重要调控网络,实时荧光定量RT-PCR与测序结果一致。【结论】本研究初步鉴定出lncRNA-ENSSSCG00000041959-RIPK1和lncRNA-ENSSSCG00000041959-IRAK1可能是影响Toll样受体信号通路的lncRNA-mRNA调控网络,为进一步探索lncRNA调控ASFV感染机体免疫反应奠定了理论基础。     

基于RNA-Seq筛选ALV-J感染汶上芦花鸡肝差异表达致病相关基因

为挖掘感染ALV-J汶上芦花鸡的肝差异表达致病相关基因。本研究以汶上芦花鸡为试验素材,分别在42和300日龄进行ALV血液病毒分离检测筛选ALV阴性和阳性个体,对ALV阳性个体有病理变化的肝组织和阴性个体肝组织进行PCR检测,分别选取3只ALV阳性（G1组）和阴性（G2组）个体的肝组织,并对G1组PCR产物测序、聚类分析确定ALV亚型,利用RNA-Seq测序技术筛选差异表达基因,并进行功能分析,利用荧光定量qRT-PCR对部分差异表达基因进行验证。结果表明,G1组样品经PCR检测、PCR扩增产物测序和聚类分析确定ALV为J亚型,转录组分析发现,共有42个差异基因在GO和KEGG中富集,其中,上调基因18个,下调基因24个。随机选取的5个差异表达基因qRT-PCR验证结果与RNA-Seq测序结果相一致。GO分析显示,差异表达基因主要涉及细胞过程、代谢过程、对刺激的反应、免疫系统等;KEGG分析显示,差异表达基因主要富集在细胞过程、信号传导、疾病、新陈代谢等信号通路。本研究通过转录组分析发现了影响ALV-J致病性的多个基因和关键信号通路,为深入了解ALV-J致病的分子机制提供了思路。     

草地早熟禾干旱胁迫转录组差异性分析

干旱是影响草地早熟禾生产力的主要因素之一。在没有参考基因组的情况下, 为了揭示草地早熟禾在干旱处理下基因表达谱的变化, 本文采用了高通量Illumina Hiseq测序平台对草地早熟禾的对照组(CK)与干旱处理组(D)进行转录组测序, 并对测序数据进行了分析研究, 进一步探究了草地早熟禾干旱应答的分子机制。结果表明, 在干旱处理下共检测到24465个差异表达的基因, 筛选后获得4143个上调基因和4415个下调基因, 共占差异表达基因总数的34.98%。经富集分析后得, 与蛋白激酶、蛋白磷酸酶、碳代谢以及ABA等相关的基因可以作为研究草地早熟禾干旱响应机制的主要研究对象。qRT-PCR分析表明, 随机选出的8个差异性表达基因的表达趋势与高通量测序结果相一致。此外, 还候选了吲哚-3-甘油磷酸合成酶、蛋白磷酸酶、已糖激酶、钙结合蛋白、叶绿素a/b结合蛋白等基因作为与草地早熟禾干旱胁迫相关的应答候选基因, 为揭示草地早熟禾耐旱分子机制奠定了基础。     

我国地方鹅种就巢性状相关转录组研究进展

随着转录组测序技术的发展,利用组学技术解析我国地方鹅就巢性的研究逐渐深入.在不同品种鹅的各个繁殖阶段,围绕下丘脑-垂体-卵巢轴的不同水平进行转录组分析,可以深入解析鹅就巢的遗传机制.文章针对转录组解析我国地方鹅繁殖性能的研究进行综述,分析不同品种鹅在各产蛋阶段的基因表达图谱,汇总与鹅繁殖性状显著相关的差异表达基因,总结...     

Combined microRNAome and transcriptome analysis of follicular phase and luteal phase in porcine ovaries

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.     

不同硒源对肝用鹅早期生长性能及免疫功能的影响

为了研究不同硒源对肝用鹅早期生长性能及免疫功能的影响,试验选用1日龄健康、体重相近的肝用鹅96只,按单因素完全随机设计分成4个组,1组为对照组,2组添加亚硒酸钠,3组添加酵母硒,4组添加纳米硒。每组3个重复,每个重复8只,公母各半。在基础日粮一致的情况下,2、3、4组分别添加同水平(0.3mg/kg)的不同硒源。试验期为9周。结果表明:各试验组体增重、采食量和料重比与对照组比较差异不显著(P0.05)。9周龄除亚硒酸钠组脾脏指数与对照组差异不显著外(P0.05),其余各试验组4周龄和9周龄免疫器官指数均显著高于对照组(P0.05),但不同硒源间差异不显著(P0.05)。4周龄和9周龄T淋巴细胞转化率亚硒酸钠组和纳米硒组均显著高于对照组(P0.05),酵母硒组显著高于亚硒酸钠组(P0.05),极显著高于对照组(P0.01)。上述结果表明,各种硒源对鹅生产性能的影响差异不显著,而对于免疫功能的影响,酵母硒优于纳米硒和亚硒酸钠。