狼山鸡下丘脑和卵巢组织全基因组DNA甲基化分析

为了获得狼山鸡性腺轴组织基因组DNA甲基化水平和模式等表观遗传信息,试验采用全基因组重亚硫酸盐测序（whole genome bisulfite sequencing,WGBS）技术检测狼山鸡下丘脑和卵巢组织基因组DNA甲基化状态,分析两组织DNA甲基化水平及特异甲基化模式。结果表明,狼山鸡下丘脑和卵巢基因组整体甲基化水平分别为4.35%和3.48%,差异显著（P<0.05）;下丘脑和卵巢中分别检测到6 150 000和10 320 000个甲基化胞嘧啶（mC）位点,其中mCG类型位点分别占69.99%和87.88%,下丘脑中非mCG位点占比约为卵巢中的2.5倍;与各染色体不同,两组织线粒体基因组中mCHH位点占比最高,其次是mCHG位点;卵巢基因组启动子区DNA甲基化水平极显著低于内含子和外显子区（P<0.01）,极显著高于基因间区（P<0.01）;下丘脑基因组启动子区DNA甲基化水平与内含子和外显子区相比差异不显著（P>0.05）,却显著高于基因间区（P<0.05）;下丘脑基因组各功能元件DNA甲基化水平均显著或极显著高于卵巢基因组（P<0.05;P<0.01）。综上,狼山鸡下丘脑和卵巢组织具有不同的DNA甲基化模式和特征,下丘脑中较高的非mCG位点比例可能在中枢神经系统发育中发挥重要作用,本研究结果为进一步分析鸡卵巢和下丘脑基因组DNA甲基化对其繁殖性能调控机制提供参考依据。     

调控香猪繁殖候选miRNA筛选

研究旨在从香猪卵巢small RNA测序数据中挖掘调控香猪繁殖的microRNA （miRNA）,解析miRNA调控香猪卵巢功能及繁殖性状的分子机制,对于猪的遗传选育具有重要意义。研究选取发情期和间情期的香猪各4头,屠宰后取其卵巢组织,提取总RNA进行small RNA测序,利用生物信息学方法检测miRNA表达谱,筛选差异表达的miRNA,预测差异表达miRNA靶基因,并对靶基因进行GO功能富集和KEGG通路富集分析。结果显示,香猪卵巢组织中miRNA在染色体上呈不均匀分布,主要分布于1号染色体和X染色体上。香猪卵巢中共有627个已知猪miRNAs表达,其中有34个差异表达miRNAs,表达量前五的分别是miR-23、let-7i-5p、miR-103、miR-30e-5p和miR-1271-5p。GO功能分析结果显示,靶基因主要参与的生物学过程是细胞过程（cellular process）,主要分布于细胞（cell）,主要分子功能是结合（binding）;KEGG显著富集通路中,促性腺激素释放激素受体通路（gonadotropin-releasing hormone receptor pathway）、胰岛素样生长因子通路（IGF pathway）和表皮生长因子受体信号通路（EGF receptor signaling pathway）与卵母细胞的发育成熟相关,因此推测miR-23b、let-7i-5p、miR-103、miR-30e-5p和miR-1271-5p可能参与了香猪繁殖调控。本研究初步筛选出5个可能调控香猪繁殖性能的miRNAs,可为从分子水平提高香猪产仔数提供理论基础。     

绵羊不同妊娠时期卵巢转录组差异表达比较分析

[目的]探究随着绵羊体内胎儿的发育,母体卵巢转录组表达水平的变化。[方法]试验绵羊为苏格兰黑面羊和特克赛尔羊F₁代杂交品种,3个妊娠时间分别为妊娠23 d、妊娠35 d和妊娠100 d。从NCBI的高通量测序数据SRA存储库下载3个妊娠时期绵羊卵巢转录组的18个样本数据。使用Z-Score方法对转录组数据进行标准化处理。将妊娠23 d、妊娠35 d的绵羊卵巢组织进行比对,妊娠35 d、妊娠100 d的绵羊卵巢组织进行比对,使用R语言的limma包进行基因差异表达分析,筛选DEGs,以P-Value<0.05和|logFC|≥2作为筛选DEGs条件。利用DAVID在线软件对DEGs进行注释及功能富集分析,并以P-Value<0.05作为信号通路显著富集标志。[结果]在妊娠23 d与妊娠35 d组绵羊卵巢中筛选出54个DEGs,妊娠35 d与妊娠100 d组绵羊卵巢中筛选出68个DEGs,两组均包含HSP90AA1、HSP90AB1、MMP2和HSP90B1基因。妊娠23 d与35 d组绵羊卵巢的54个DEGs显著富集到包括内质网蛋白质加工信号通路、核糖体信号通路、雌激素信号通路、癌症蛋白聚糖信号通路、扩张型心肌病信号通路、肥厚型心肌病信号通路6条通路;妊娠35 d、妊娠100 d组绵羊卵巢的68个DEGs显著富集到包括癌症相关信号通路、黏着斑信号通路、抗原加工与提呈信号通路、细胞外基质受体相互作用信号通路、细菌侵袭上皮细胞信号通路、内质网蛋白质加工信号通路、核糖体信号通路、雌激素信号通路、PI3K-Akt信号通路、癌症蛋白聚糖信号通路和前列腺癌信号通路11条通路。[结论]在妊娠期间雌激素信号通路上的HSP90AA1、HSP90AB1、MMP2和HSP90B1基因表达有很大变化,这些基因可能对绵羊妊娠维持有着重要的作用。     

Porcine reproductive and respiratory syndrome virus whole-genome sequencing efficacy with field clinical samples using a poly(A)-tail viral genome purification method

The genomic surveillance of porcine reproductive and respiratory syndrome virus (PRRSV) is based on sequencing of the ORF5 gene of the virus, which covers only 4% of the entire viral genome. It is expected that PRRSV whole-genome sequencing (WGS) will improve PRRSV genomic data and allow better understanding of clinical discrepancies observed in the field when using ORF5 sequencing. Our main objective was to implement an efficient method for WGS of PRRSV from clinical samples. The viral genome was purified using a poly(A)-tail viral genome purification method and sequenced using Illumina technology. We tested 149 PRRSV-positive samples: 80 sera, 33 lungs, 33 pools of tissues, 2 oral fluids, and 1 processing fluid (i.e., castration liquid). Overall, WGS of 67.1% of PRRSV-positive cases was successful. The viral load, in particular for tissues, had a major impact on the PRRSV WGS success rate. Serum was the most efficient type of sample to conduct PRRSV WGS poly(A)-tail assays, with a success rate of 76.3%, and this result can be explained by improved sequencing reads dispersion matching throughout the entire viral genome. WGS was unsuccessful for all pools of tissue and lung samples with Cq values > 26.5, whereas it could still be successful with sera at Cq ≤ 34.1. Evaluation of results of highly qualified personnel confirmed that laboratory skills could affect PRRSV WGS efficiency. Oral fluid samples seem very promising and merit further investigation because, with only 2 samples of low viral load (Cq = 28.8, 32.8), PRRSV WGS was successful.     

Interspecific hybrids betweenAllium fistulosumandAllium schoenoprasumreveal carotene-rich phenotype

Interspecific hybridization is an effective method to generate a new crop that gains available functions in a short time. Interspecific hybrids (2n= 16) betweenAllium fistulosumL. (2n= 16) andAllium schoenoprasumL. (2n= 16) were produced by reciprocal crossings through ovary culture, but the hybrids were much fewer in the combination usingA. schoenoprasumas a seed plant. All the hybrids have eight long chromosomes originated fromA.fistulosumand eight short chromosomes originated fromA. schoenoprasum. In addition, the hybridity was confirmed by randomly amplified polymorphic DNA (RAPD) analysis and cleaved amplified polymorphic sequence (CAPS) analysis of the rDNA internal transcribed spacer (ITS) region. The interspecific hybrids showed a vigorous growth habit; their foliage was slightly bloomy and deep green. The hybrids did not form bulbs, but rather propagated vegetatively by tillering. Carotene contents of the hybrids and both parents were quantified using high-performance liquid chromatography (HPLC). The contents of all edible parts of the hybrids were approximately seven times higher than those of either parent. These results indicate that the hybrid is a new and carotene-rich vegetable ofAlliumspecies.     

Optimization of culture conditions for improved plant regeneration efficiency from wheat microspore culture

The production of haploid plants through microspore culture is a very important tool for plant breeding. However, progress in microspore culture for many species has been hampered by a number of factors that have resulted in low recovery of regenerated green plants. In this study, a series of experiments were conducted to increase the regeneration of haploid green plants from isolated wheat microspores. The use of different basal media and variations in media components resulted in the increased recovery (approximately double) of regenerated haploid wheat plants. Our findings demonstrate that CHB medium, in combination with 2,4-d, was a better medium for embryoids induction and plant regeneration than medium MC17 with either 2,4-d or PAA growth hormones. Wheat microspores cultured without ovary co-cultivation did not respond. Furthermore, high efficiency of microspore derived embryoids (up to 296 MDEs per 100 anthers) and green plant regeneration (up to 71 green plants per 100 anthers) were achieved by the use of gelrite instead of agarose as a gelling agent, and by the addition of media additives such as spent medium or MET.     

福州地区甘蔗黄叶病病原分子鉴定及电镜检测

电镜观察表明，感病甘蔗叶片的韧皮部伴胞内存在着大量类似甘蔗黄叶病毒（SCYLV）的病毒粒子；利用RT-PCR扩增出556 bp的目的片段，氨基酸和核苷酸序列分析表明，该片段是SCYLV外壳蛋白基因的一部分，与其他国家SCYLV分离物同源性达95%以上；应用组织印迹杂交免疫检测技术，在YLS病症的叶片中脉韧皮部出现紫红色斑块，呈阳性反应。综合分析认为福州地区甘蔗黄叶病的病原体为SCYLV。     

Distribution of antibodies against porcine circovirus type-2 (PCV2) in single site and multi-site farrow-to-finish farms in Brazil

A comparative serologic study was performed in seven single site (SS) farrow-to-finish farms and four multi-site (MS) farrow-to-finish farms, with or without post-weaning multisystemic wasting syndrome (PMWS). In each farm, 30 blood samples were collected for each category of the production cycle: sows, farrowing crate, nursery, grower pigs, and finishing pigs. Sera were evaluated for the presence of antibodies to porcine circovirus type-2 (PCV2) via immunoperoxidase monolayer assay. Serologic profiles for PCV2 were different between SS and MS farms. Seroconversion following the decline in maternal antibodies occurred at a later stage on SS farms (grower pigs) than MS farms (nursery pigs). MS farms tended to have lower antibody titers than SS farms in the categories of sow, piglet, and nursery, while higher antibody titers were found in grower pigs. Characterization of serologic profiles for different farms may provide important information for the adoption of vaccination programs.     

PRRSV感染猪体内多杀性巴氏杆菌的分离与血清型鉴定

应用常规方法和PCR技术,对来自安徽肥西、庐江、肥东、定远、桐城5个地区猪场检测为猪繁殖与呼吸综合征病毒(PRRSV)阳性的肺脏进行多杀性巴氏杆菌(Pm)的分离及其血清型鉴定。结果显示,49份病料中分离鉴定出7株Pm,且均属于A型,分离率为14.3%,高于其他细菌(副猪嗜血杆菌、猪链球菌)的检出率。表明猪繁殖与呼吸综合征(PRRS)患猪继发或混合感染Pm的比率较高,而且A型Pm是安徽省感染猪群中的主要血清型。     

Regulation of Insulin Action by Diet and Exercise

The regulation of the cellular actions of the hormone insulin is essential to the maintenance of macronutrient metabolism, body weight regulation, and a surprisingly diverse range of other integrative physiologic functions. Because of the diverse targets of insulin action, any dysfunction in insulin is likely to have systemic consequences. Although type 1 and type 2 diabetes are the most obvious clinical consequences of impaired insulin synthesis and insulin action, respectively, there are also subclinical disorders that attend defects in the function of insulin. In humans and horses, the “metabolic syndrome” is characterized by a cluster of metabolic sequelae that arise as a result of insulin resistance. Importantly, both diet and exercise can regulate insulin action and can thus be leveraged as treatment tools to prevent and treat the metabolic syndrome. The aim of this review is to characterize the integrative biology of insulin action and to describe the role of diet and exercise in regulating tissue responsiveness to insulin.