绵羊Izumo1基因多态性及其与产羔数关联分析

为研究Izumo1基因多态性及与产羔数关系,本研究利用PCR和直接测序法对小尾寒羊和苏尼特羊的Izumo1基因DNA序列进行扩增、测序和BLAST分析,并和本课题组前期绵羊重测序数据进行比对,筛选出Izumo1基因SNP位点。同时,采用Sequenom MassARRAY~?技术进行基因分型,并对其SNP位点的基因型和等位基因频率在各群体中的分布进行研究。结果表明:小尾寒羊Izumo1基因DNA全长序列3 385 bp,苏尼特羊Izumo1基因DNA全长序列3 382 bp;筛选出8个Izumo1基因SNP位点,经过初步筛选,将在小尾寒羊、苏尼特羊、滩羊、萨福克羊、杜泊羊、草原藏羊这6个绵羊品种中位点分布没有差异的SNP位点排除,最终筛选获得的g.54412135AG和g.54412107CA 2个位点。Izumo1基因g.54412135AG位点在多羔和单羔绵羊品种中存在GG、GA和AA 3种基因型,G基因为优势等位基因;g.54412107CA在多羔品种中存在CC和CA 2种基因型,而在单羔品种中存在CC、CA和AA 3种基因型,C基因为优势等位基因,其基因型频率和基因频率在单、多羔绵羊群体间的分布差异均极显著(P0.01);群体遗传学分析得出g.54412135AG多态位点在6个品种中的多态信息含量(PIC)都属于低度多态(PIC0.25);g.54412107CA多态位点在杜泊羊中属于中度多态(0.25PIC0.5),在其他5个品种中都属于低度多态(PIC0.25),然而,关联分析发现Izumo1基因g.54412135AG和g.54412107CA位点的不同基因型与小尾寒羊不同胎次产羔数之间不存在显著关联(P0.05)。     

Effect of melatonin on visceral fat deposition,lipid metabolism and hepatic lipo-metabolic gene expression in male rats

Melatonin (MT) influences lipid metabolism in animals; however, the mechanistic effect of melatonin on liver fat and abdominal adipose deposition requires further clarity. In order to study the effects of melatonin on lipid metabolism, and hepatic fat and abdominal adipose deposition in animals, twenty Sprague–Dawley (SD) rats of 6 weeks of age with similar bodyweight were randomly divided into two groups: control (CTL) and MT-treated (10 mg/kg/day). During a 60-day experiment, food intake and bodyweight were measured daily and weekly respectively. At the end of treatment, blood samples were collected to collect plasma to quantify hormones and metabolic indicators of lipid metabolism. In addition, organ and abdominal adipose depots including liver, and omental, perirenal, and epididymal fat were weighed. Liver tissue was sampled for sectioning, long-chain fatty acid (LCFA) quantification, and gene chip and Real-time quantitative PCR (qPCR) analyses. The results showed that liver weight and index (ratio of liver weight to body weight) in MT group reduced by 20.69% and 9.63% respectively; omentum weight and index reduced by 59.88% and 54.93% respectively, and epididymal fat weight reduced by 45.34% (p = 0.049), relative to CTL. Plasma lipid indices, triglyceride (TG), high-density lipoprotein (HDL), low-density lipoprotein (LDL) and total cholesterol (TC) with MT treatment decreased significantly compared with the control. Fat and 8 LCFA content in liver in MT group also decreased. Gene chip and qPCR demonstrated that there were 289 genes up-regulated and 293 genes down-regulated by MT. Further analysis found that the mRNA expression of lipolysis-related genes increased, while the mRNA expression of lipogenesis-related enzymes decreased (p < 0.05) with MT. This study concluded that melatonin greatly affected fat deposition, and hepatic LCFA supply and the expression of genes associated with lipogenesis and lipolysis.     

羊口疮病毒安徽株116基因的生物信息学分析及原核表达

为了对羊口疮病毒（Orf virus, ORFV）安徽株116基因进行生物信息学分析及原核表达。以ORFVAH-F10株为模板PCR扩增116基因，使用生物信息学软件预测该基因编码蛋白的结构与功能。同时将扩增产物克隆至pET-32a（＋）原核表达载体，经BamH I及EcoR I双酶切鉴定后进行测序，构建pET-32a（＋）-116重组质粒。转化至大肠杆菌Rosetta（DE3）进行IPTG诱导，纯化蛋白后免疫BALB/c雌性小鼠制备多克隆抗体。测序结果显示，ORFV116基因全长561 bp，编码186个氨基酸。生物信息学分析结果显示：该蛋白分子量约为20.33 kDa，为不稳定亲水性蛋白；无信号肽、保守结构域及跨膜结构域；含有2个N糖基化位点和42个磷酸化位点；二级结构以无规则卷曲为主，分别为延伸链（5.91%）、α-螺旋（14.52%）与无规则卷曲（79.57%）；预测该蛋白含有20个可能的B细胞优势抗原表位与4个CTL细胞表位。SDS-PAGE及Western-blot结果显示，ORFV116蛋白大小约为51 kDa，主要以可溶形式表达且具有良好的反应原性。     

Alteration of the haemocyte parameters,expression of immune‐related and neurohormone bursicon genes of giant river prawn Macrobrachium rosenbergii (de Man) in the response to salinity stress and ammonia stress

We examined the effects of salinity stress and ammonia stress on alteration of the haemocyte count, phenoloxidase activity, expressions of immune‐related genes including prophenoloxidase (proPO), crustin, heat shock protein 70 (HSP70), and expressions of stress‐responsive neurohormone (Bur‐α and Bur‐β) in the thoracic and abdominal ganglia of giant river prawn Macrobrachium. These parameters of prawn that subjected to salinity stress (transferred from 0‰ to 5‰ and 10‰), and subjected to ammonia‐nitrogen (ammonia‐N) stress (transferred from 0 to 0.262 mg/L and 0.786 mg/L) were examined after 0, 3, 6, 24, 72 and 168 hr respectively. During the initial period of 3, 6 and 24 hr, granulocyte haemocyte (granular and semi‐granular hemocyte) count and PO activity significantly decreased, while expressions of Bur‐α and Bur‐β significantly increased. After 24 hr, granulocyte haemocyte count and PO activity significantly increased, whereas expressions of Bur‐α and Bur‐β significantly decreased. The expressions of proPO, crustin and HSP70 were significantly downregulated in the prawn that subjected to salinity stress and ammonia‐N stress at all time periods of 3–168 hr. In conclusion, changes in the granulocyte haemocyte count of M. rosenbergii following salinity stress and ammonia‐N stress are closely associated with the changes of Bur‐α and Bur‐β expressions.     

砂藓(Racomitrium canescens)RcRab基因的克隆及生物信息学分析

本研究以砂藓(Racomitrium canescens)为材料,从砂藓干旱处理转录组数据库中筛选获得小G结合蛋白基因Rab的mRNA序列,通过RT-PCR法扩增,得到RcRab基因的CDS序列,并对其进行了生物信息学分析。结果表明,该基因c DNA全长为1171 bp,包含636 bp的开放阅读框,编码211个氨基酸。预测该基因有Rab家族的保守结构域-G结构域。RcRab蛋白分子质量为23.29 kD,理论等电点为5.76,具有亲水性,不属于跨膜蛋白且不存在信号肽。进化树分析表明,RcRab蛋白质与小立碗藓(Physcomitrella patens)Rab蛋白质亲缘关系最近。本研究可为进一步探索RcRab基因在苔藓植物中的抗旱机制提供数据基础。     

罗氏沼虾细胞色素P450家族CYP302a1基因克隆及其在蜕皮周期中的表达

蜕皮是甲壳动物重要的生理活动,与其蜕皮激素的合成密切相关,细胞色素P450(CYP)302a1是甲壳动物蜕皮激素合成通路中的关键酶之一。本研究克隆了罗氏沼虾CYP302a1基因(Mr-CYP302a1),cDNA全长1859 bp,开放阅读框(ORF)为1629 bp,编码543个氨基酸(aa),分子量大小为61.09 ku,等电点为8.42。氨基酸序列分析显示CYP302a1基因的保守结构域含有5个P450基因家族特征保守区域:heme-binding、helix-K、helixC、helix-I及PERF。系统进化分析结果显示Mr-CYP302a1首先与绿虾CYP302a1聚为一支,然后与凡纳滨对虾及三疣梭子蟹等十足目甲壳动物的CYP302a1聚为一支,与甲壳动物的亲缘关系最近。实时荧光定量PCR(qRT-PCR)检测表明Mr-CYP302a1在罗氏沼虾的多个组织中均有表达,其中在Y器官中的表达量最高,性腺中次之。同时研究发现,MrCYP302a1基因在罗氏沼虾的蜕皮后期(A期和B期)表达量很低,蜕皮间期(C期)表达量开始上升,在蜕皮前期D1亚期达到峰值。对Mr-CYP302a1进行蛋白表达及多克隆抗体制备,蛋白印迹法(Western blot,WB)检测表明Mr-CYP302a1蛋白在罗氏沼虾Y器官中的表达量最高,在蜕皮过程中的蜕皮前期D1亚期达到峰值。综上所述,该基因在罗氏沼虾的蜕皮过程中扮演着十分重要的角色。     

2株可培养细菌氮代谢特性的研究

研究旨在分析土壤中可培养细菌菌株的氮代谢特征,并进一步探讨微生物在土壤氮素转化中的可能作用机制。以2株分离自苹果园土壤的细菌菌株SY5-4和SY11-10为试材,采用传统培养方法结合分子检测技术,分别测定菌株生长特性及其氮素转化能力。研究结果表明,异养条件下,菌株SY5-4和SY11-10的世代时间分别为243.5 min和202.7 min。菌株生长过程中,培养液中铵态氮浓度始终维持在较高水平,铵态氮、亚硝态氮和硝态氮浓度均表现出先升后降的趋势。硝化（amoA和hao）和反硝化（nosZ、norB、nirK和nap）基因检测结果表明,菌株SY11-10具有多种氮素转化潜能。综上,供试菌株培养过程中,培养液中氮素发生变化,并在菌体中检测到不同氮转化基因,表明菌株参与多种氮代谢途径。     

基于SNP标记的爆裂玉米农家品种遗传多样性

为了评价广西爆裂玉米农家品种的遗传多样性,利用56K SNP芯片技术对中国广西45个爆裂玉米农家品种、中国2个爆裂玉米杂交种和6个南美爆裂玉米种质进行全基因组扫描,获得多态性标记14 338个,基因分型为6 种类型。其中A/G类型最多,为5 805个;A/T类型最少,为149个。1号染色体的多态性SNP位点最多,为2 302个;10号染色体最少,为848个。45个农家品种间平均遗传相似系数为0.62,变幅为0.41~0.99,总体遗传相似度高。聚类分析将参试品种划分为三大类群,相同来源的农家品种大多聚在一起;主成分分析显示大部分农家品种聚在一起,与杂交品种和南美品种之间没有交集,表明农家品种遗传相似性较高,与杂交品种和南美品种遗传差异较大。     

Intermediary metabolic response and gene transcription modulation on the Sub-Antarctic notothenioid Eleginops maclovinus (Valenciennes, 1930) injected with two strains of Piscirickettsia salmonis

Pathogen interactions with cultured fish populations are well studied, but their effects on native fishes have not been characterized. In Chile, the disease caused by bacterial species Piscirickettsia salmonis represents one of the main issues and is considered to be one of the important pathogens in the field of aquaculture. They have been found to infect native fish. Therefore, it is necessary to understand the impact of P. salmonis on native species of local commercial value, as well as the potential impact associated with the emergence of antibiotic-resistant strains of P. salmonis. Due to this purpose, the native fish Eleginops maclovinus was used in our study. Fish were randomly distributed in tanks and intraperitoneally inoculated with two strains of P. salmonis. No mortality was recorded during the experiment. Cortisol, glucose and total α-amino acid levels increased in fish injected with AUSTRAL-005 strain compared to sham-injected and LF-89-inoculated fish. Moreover, results showed an increase in the activity of carbohydrates and lipids metabolism in liver; and an increase in the carbohydrates, lipids and total α-amino acid metabolism in muscle after injection with AUSTRAL-005. Our results suggest that P. salmonis modulates the physiology of E. maclovinus and the physiological impact increase in the presence of the antibiotic-resistant strain AUSTRAL-005.     

Koi herpesvirus and carp edema virus threaten common carp aquaculture in Croatia

Common carp (Cyprinus carpio) is a very important fish species for warm-water aquaculture in Croatia. All Croatian carp farms are subjected to a surveillance programme for the presence of koi herpesvirus (KHV), causing a deadly disease called koi herpesvirus disease (KHVD). However, there is no surveillance for other viral pathogens of importance like carp edema virus (CEV), a causative agent of koi sleepy disease (KSD). During regular testing within the KHVD surveillance programme, we tested samples for CEV simultaneously. The screening indicated possible outbreaks of KHVD and KSD. During 2016, KHVD broke out in an isolated area and soon thereafter a KHV eradication programme was successfully performed. However, during 2018 and 2019, two additional mortality events occurred in lakes in the southern part of Croatia during the spring. Samples from both events tested positive for CEV. An epidemiological investigation confirmed the introduction of infected carps from an infected farm to one of the lakes. To prevent the spreading of CEV into open waters, it is of utmost importance to introduce CEV testing before fish movement or to perform regular testing of all carp farms in the country to determine CEV prevalence for the purpose of implementation of control measures.