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利用靶基因步行 PCR 方法扩增得到鸭肠炎病毒(DEV Clone-03)5352 bp 基因片段,并进行克隆和测序.序列分析发现,该片段包含2个ORF.Blast 分析表明,这2个ORF分别与单纯疱疹病毒1型(HSV-1)UL51 和UL52 基因同源,命名为DEV Clone-03 UL51和UL52,分别编码252和1124个氨基酸.DEV Clone-03 UL51和UL52基因排列方式与HSV-1同源基因相一致,为"头对头"的转录方向,两个ORF间有236 bp 间隔序列.用DNAStar(Megalign)软件分别对这2个ORF和11株α-疱疹病毒参考毒株进行分析,发现DEV Clone-03 UL51 与HSV-1的氨基酸序列有相对较高同源性,DEV Clone-03 UL52与EHV-4 同源性相对较高.并且DEV Clone-03UL51基因在α-疱疹病毒中比较保守.与HSV-1 UL52 蛋白相似,DEV Clone-03 UL52蛋白C末端存在锌指结构.系统发育分析表明,DEV Clone-03与α疱疹病毒亚科成员的关系比较保守,与Mardivims属的成员具有较近的亲缘关系. 相似文献
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鸡传染性喉气管炎病毒TK基因的克隆与鉴定 总被引：2，自引：0，他引：2
根据鸡传染性喉气管炎病毒（ILTV）TK基因的核苷酸序列，设计并合成了1对引物，利用该引物扩增出了长1800bp的TK基因核苷酸片段。将该基因克隆入PGEM-T载体后，对其进行酶切分析和序列测定。结果表明，扩增片段与发表的ILTV的TK基因核苷酸序列一致。以ILTV、正常绒毛尿囊膜组织、鸡传染性支气管炎病毒（IBV）、鸡新城疫病毒（NDV）感染鸡胚的尿囊膜DNA为模板进行PCR反应，结果该PCR反应体系对ILTV是特异的。敏感性试验表明，该PCR系统能检出50pg的ILTV感染尿囊膜的DNA。 相似文献
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商品肉鸭鸭瘟病毒的分离与鉴定 总被引：1，自引：0，他引：1
采用鸭胚成纤维细胞培养从山东和北京两地暴发的鸭瘟临床病例中分离到两株鸭肠炎病毒（DEV），分别命名为SD和BJ。以单抗介导的间接免疫荧光（IFA）检测方法，对两个分离株感染细胞滴片进行间接IFA检测，可见感染细胞内有明显的蓝绿色荧光。试验感染7日龄北京鸭可引起鸭瘟的典型临床症状．死亡率为100％（3／3），取试验感染死亡鸭肝脏、法氏囊和脑组织等制备石蜡包埋切片，利用单抗进行免疫组化检验，除脑组织外均检测到病毒抗原。根据在GenBank上已发表的DEV两段序列设计两对引物，采用聚合酶链式反应（PCR）对野毒sD株人工感染鸭肝脏和BJ珠自然发病鸭肝脏病科提取核酸为模板进行扩增，得到预期大小为765bp和1954bp的目的片段，对长片段进行测序，与发表序列进行比较，毒株间的碱基序列同源性达到99．73％。 相似文献
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为了研究鸭肠炎病毒（Duck enteritis virus,DEV）强毒株在感染鸭体内的动态分布规律,根据DEV的gD基因序列设计检测引物RT-gDF和RT-gDR,建立了特异性强、敏感性高、重复性好的检测DEV的SYBR GreenⅠ实时荧光定量PCR方法。应用已建立的方法,对DEV强毒参考株感染鸭的组织脏器进行了定量检测。结果显示,接种后6 h即可在所有受检组织中检测到DEV DNA,随着病程发展,DNA拷贝数持续升高,直至接种5 d后感染鸭全部死亡。其中结肠病毒DNA拷贝数为最高,达到1013.26 copies,法氏囊、肝脏和盲肠次之,约1012 copies。本研究证实了DEV强毒对感染鸭的免疫器官、神经组织和消化系统均具有广泛的嗜性。 相似文献
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鸡胚成纤维细胞培养鸭瘟病毒的试验 总被引：1，自引：0，他引：1
将鸭瘟病毒强毒（DPV34）经12日龄鸭胚连续传2代，收获的尿囊液DPV34F2作为细胞培养的病毒接种于鸡胚成纤维细胞，连续传代5次。结果，从第2代开始，接种DPV的鸡胚成纤维细胞出现细胞病变，随首代次的增加，细胞病变愈加明显。经电镜观察，第5代细胞培养物中有典型的DPV病毒粒子；将第5代培养物接种鸭胚，出现典型鸭瘟病变；用PCR方法检测培养物，也出现DPV的特征DNA带。 相似文献
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鹅细小病毒主要结构蛋白VP3基因的克隆与序列分析 总被引：6，自引：1，他引：5
将GPV H2分离株接种于13日龄非免疫鸭胚，收集接毒后3－7日死亡鸭胚的尿囊液。纯化病毒，通过PCR技术，从病毒基因组DNA中扩增出病毒衣壳蛋白VP3完整基因片段，经酶切鉴定后直接与pMD 18-T质粒载体连接，转化感受态大肠杆菌TG1。提取重组质粒经PCR鉴定和酶切鉴定后，对插入片段进行序列测定及分析。结果表明；鹅细小病毒H1分离株VP3基因全长1605bp,编码534个氨基酸，与国外已发表的鹅细小病毒B株核苷酸序列同源性为98．5％，氨基酸序列同源性为98．3％，表明这二个毒株亲缘关系相近。 相似文献
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《中国预防兽医学报》2015,(5)
为鉴定鸭肠炎病毒(DEV)的非必需基因,本研究利用增强型绿色蛋白(EGFP)基因为报告基因,通过在转染的鸡胚成纤维细胞(CEF)中同源重组,将其插入DEV基因组的TK基因中,经筛选和纯化绿色荧光病毒蚀斑,获得表达EGFP的重组DEV,命名为r DEVTK-EGFP。对重组病毒与亲本病毒DEV Clone-03复制动力学比较结果显示,重组病毒滴度显著低于亲本病毒(p0.01)。对重组病毒在复制过程中感染细胞能力和荧光表达时相检测结果表明,在感染后48 h~72 h表达荧光信号细胞明显增多,该结果与重组病毒复制动力学一致。将重组病毒在CEF中连续传代至20代,仍具有遗传稳定性。该研究结果表明,TK基因为DEV复制非必需基因,可以作为外源基因插入位点,用于禽用新型基因工程疫苗的研制。 相似文献
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应用葡萄球菌A蛋白协同凝集试验快速检测鸭肝炎病毒的研究 总被引：9，自引：2，他引：7
用提纯的鸭肝炎病毒（ＤＨＶ）免疫家兔制备高免血清，常规方法提取ＩｇＧ，与１％葡萄球菌Ａ蛋白（ＳＰＡ）阳性菌悬液混合，以ＰＨ７．２０．２ｍｏｌ／ＬＰＢＳ作缓冲液，制成鸭病毒性肝炎（ＤＶＨ）ＳＰＡ协同凝集试验（ＳＰＡ－ＣｏＡ）诊断试剂，同时制备阴性对照试剂。用ＳＰＡ－ＣｏＡ检测人工感染致死的雏鸭肝脏、含ＤＨＶ强毒的鸭胚液、适应鸡胚的疫苗毒鸡胚尿囊毒的检出率均为１００％。用ＳＰＡ－ＣｏＡ检测强毒时，样品 相似文献
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魏静 《四川畜牧兽医学院学报》2009,(4):28-32
在现代法律秩序中,商会自治规范是制定法的基础和必要的补充,甚至在某些方面替代了制定法;商会自治规范主要包括商会组织规范、行为规范、惩罚规范以及争端解决规范等;其效力仅及于其内部成员;商会自治规范和制定法之间存在冲突,但也存在整合的基础。 相似文献
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本文概述了猪的毛色类型、猪的毛色遗传模式,着重综述了猪毛色基因分子基础的研究进展,指出存在问题并就未来发展方向做了思考。 相似文献
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以国际标准强毒Ｒ株人工感染非免疫产蛋鸡,定时扑杀,分别从鼻窦、眶下孔、气管、肺、气囊、卵巢和输卵管分离ＭＧ,并收集感染鸡所产蛋分离ＭＧ。结果表明,人工感染４８小时后上、下呼吸道及肺已被全面感染,９６小时气囊已被感染,１２０小时输卵管已能分离到ＭＧ,卵巢始终分离不到ＭＧ。人工感染鸡自１４４小时便能在其所产蛋中分离出ＭＧ。药物治疗能在７２小时内消除感染,油乳剂苗则需２４天后逐渐降低蛋内ＭＧ分离率,药物卵内注射、种蛋药浴、高温处理均能杀死卵内ＭＧ,但以研制的种蛋浸泡剂药浴效果为最好。 相似文献
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REASONS FOR PERFORMING STUDY: Centesis of the bicipital bursa using an 8.9 cm long spinal needle has been reported but the alternative of employing a 3.8 cm long hypodermic needle requires validation. OBJECTIVE: To compare the efficacy of 2 different methods of centesis of the bicipital bursa and to evaluate the usefulness of ultrasonographic imaging to determine the location of solution administered when centesis of the bursa is attempted. METHODS: For Trial 1, 6 clinicians, who had no previous experience of centesis of the bicipital bursa, attempted to inject a solution composed of an aqueous radiopaque contrast medium and physiological saline solution (PSS) into the bicipital bursae of 2/12 horses using the previously described distal approach to inject one bursa and a proximal approach to inject the contralateral bursa. The bicipital tendon and bursa were examined ultrasonographically before and after injection; and both shoulders were examined radiographically to identify the location of the medium. In Trial 2, another 6 clinicians, also with no previous experience of centesis, repeated Trial 1, using 6 horses, but the radiopaque contrast medium was mixed with air instead of PSS. RESULTS: Accuracy of centesis using the proximal approach was 39% and that of the distal approach 28%. Ultrasonographic examination of the shoulder allowed the location of solution and air to be accurately predicted in all 12 shoulders examined. CONCLUSIONS: Clinicians who have had no previous experience performing centesis of the bicipital bursa are unlikely to be successful in centesis using either approach. Radiographic examination after injecting a radiopaque contrast medium may be necessary to assess the success of centesis especially if bursal fluid is not obtained during centesis. Injecting air along with the radiopaque contrast medium provides more accurate ultrasonographic confirmation of centesis and better radiographic definition than does injection without air. 相似文献
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Effects of size of ingestively masticated fragments of plant tissues on kinetics of digestion of NDF
Ingestively masticated fragments were collected and sized via sieving. Different sizes of esophageal masticate and ruminal digesta fragments, and ground fragments of larger masticated pieces were incubated in vitro, and undigested NDF remaining at intervals of up to 168 h of incubation was determined. The ruminal age-dependent time delay (tau) for onset of digestion of NDF was positively correlated (P < 0.004) with the mean sieve aperture estimated to retain 50% of the fragments between successive sieve apertures (MRA). Degradation rate of potentially degradable NDF (PDF) and level of indigestible NDF were not related (P > 0.10) to MRA of masticated and ground fragments. Estimates of tau were positively related to MRA, with slopes of bermudagrass < corn silage < ruminal fragments of corn silage. It was concluded that fragment size-, and consequently, ruminal age-dependent onset of PDF degradation of a mixture of different fragment sizes results in an age-dependent rate of degradation of the more rapidly degrading of two subentities of PDF. Models are proposed that assume a tau before onset of simultaneous degradation of PDF from two pools characterized as having gamma-modeled age-dependency and age-constant rates. The ruminal age-dependent pool seems to be associated with the faster-degrading pool, and its rate parameter increases with range in MRA in the population of fragments. Conceptually, the ruminal age-dependent rate parameter for PDF degradation seems to represent a composite of several effects: 1) effects of the size-dependent tau; 2) range in MRA of the population of ingestively masticated fragments; and 3) subentities of PDF that degrade via more rapid age-dependent rates compared with subentities of PDF that degrade via age-constant rates. The estimated fractional rates of ruminative comminution of ingestively masticated fragments (0.060 to 0.075/h) were of a magnitude similar to the mean fractional rates of PDF digestion (0.030 to 0.085/h), which implies that ruminative comminution may be first-limiting to fractional rate of PDF digestion. The in vivo roles of ingestive and ruminative mastication of fragments on PDF degradation must be considered in any kinetic system for estimating PDF digestion in the rumen. These results and others in the literature suggest that the rate of surface area exposure rather than intrinsic chemical attributes of PDF may be first-limiting to degradation rate of PDF in vivo. 相似文献