共查询到19条相似文献，搜索用时 125 毫秒
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鉴别牛早期胚胎性别PCR方法引物的设计与筛选 总被引：6，自引：2，他引：6
根据牛Y-染色体特异重复序列、睾丸特异蛋白基因以及性别决定基因序列设计合成5对公牛Y-染色体特异引物，依据牛骨胳肌α肌动蛋白前体基因和微卫星DNA序列设计合成4对牛DNA特异引物(内标引物)。单重PcR扩增牛基因组DNA，筛选出4对牛Y-染色体特异引物和1对牛DNA特异内标引物。将不同的Y-染色体特异引物与内标引物组合，多重PCR扩增牛基因组DNA、已知性别的牛成纤维细胞和克隆胚胎，筛选出2个可用于牛早期胚胎性别鉴别的PCR引物组合：B34／A12和B78／A12。 相似文献
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结缕草属植物耐盐性SRAP分子标记研究 总被引：10，自引：5，他引：5
本研究应用混合集群分析法,通过建立结缕草属植物耐盐性两极端类型材料DNA池对400对SRAP引物组合进行筛选,得到111对多态性引物组合,再以日本结缕草耐盐两极端材料DNA对111对具有特异带的引物组合进行进一步筛选,从中获得22对多态性引物组合,最后用群体各单株对具有耐盐特异带的引物组合进行验证,获得了7个与结缕草属植物耐盐性紧密相关的SRAP分子标记,依据扩增条带的大小分别命名为:Me9-Em4260、Me9-Em18720、Me13-Em18500、Me5-Em20180、Me14-Em7220、Me6-Em17600、Me2-Em18260,以上筛选得到的SRAP分子标记将为深入开展结缕草属植物分子标记辅助育种及耐盐基因克隆奠定基础。 相似文献
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通过对GenBank中注册的CSFV序列进行比对分析,针对CSFV NPro/C基因中的保守区域,设计合成了一套RT-LAMP引物,应用含有Npro/C基因的阳性质粒株对RT-LAMP引物进行验证.在成功扩增出特异片段的基础上,用CSFV RNA优化RT-LAMP反应体系和条件,建立了CSFV的可视化RT-LAMP快速... 相似文献
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甘肃省小麦条锈病主要流行小种的RAPD分析及Su11-4小种SCAR标记建立 总被引：1，自引：1，他引：0
本研究应用RAPD标记技术对甘肃省小麦条锈菌主要流行的8个生理小种进行多态性标记分析,旨在寻找小麦条锈菌不同生理小种的特异性标记,共选用220条10碱基随机引物进行筛选,其中有147条可得到稳定清晰的扩增条带。研究结果显示,通过使用147条引物对甘肃省流行的8个条锈菌的生理小种进行RAPD分析,发现各致病小种间遗传变异丰富,其中引物S301在条中33号中扩增得到约507bp的特异条带;引物S39在条中32扩增得到长度约183bp的特异条带;引物S36在Hybrid46-8扩增得到约510bp的特异条带;引物S2140在Su11-4扩增得到约317bp的特异条带。另外,本研究还对扩增得到的特异片段进行回收并进行测序分析,其中依据小麦条锈菌生理小种Su11-4特异条带的测序结果设计特异引物,成功将其转化为对Su11-4小种特异的SCAR标记,这对不同条锈菌生理小种的快速准确检测具有重要意义。 相似文献
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为了快速检测牛传染性鼻气管炎,采用SDS-蛋白酶K法,提取病毒模板DNA。根据IBRV gB基因序列设计了1对特异引物,在其上下游引物的内侧又分别设计了1对引物,以这4条引物对IBRV模板进行扩增。结果成功扩增出预期目的片段,建立了巢式PCR检测方法。敏感性、特异性等检测试验结果表明该方法能特异检测IBR病毒。本方法具有快速、灵敏、特异的优点,适用于在牛及其遗传物质的进出口检疫中进行牛传染性鼻气管炎快速病原鉴定。 相似文献
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本研究以圭亚那柱花草‘1979’（Stylosanthes guianensis ‘1979’,母本,花粉不育）和‘热研2号’柱花草（轮回父本）的BC1F1代群体为材料,利用简单序列重复区间扩增多态性（Inter-simple Sequence Repeat,ISSR）技术,结合集群分组分析法（Bulked Segregate Analysis,BSA）构建花粉育性基因池。结果表明：利用100条ISSR引物筛选出在不育基因池中有特异条带的引物共11条;经BC1F1群体单株验证,有8个标记仅在花粉不育个体中出现,表明其与花粉不育基因连锁;将这8个特异性片段回收、克隆、测序,并分别设计8对特异序列特异扩增区域（Sequenced Charaeterized Amplified Region,SCAR）引物,对BC1F1单株进行验证,发现这8条片段仅在不育个体中扩增出目标条带,表明8 ISSR特异片段成功转化为8个SCAR标记。本研究结果为建立柱花草稳定的雄性不育系提供新的理论基础,为柱花草杂交育种奠定基础。 相似文献
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根据GenBank中鸡传染性法氏囊病病毒(IBDV)A片段保守区域设计、合成了一对特异性引物(RP1/RP3),以期建立特异、灵敏的IBDVRT-PCR检测方法。结果显示RP1/RP3具有良好的特异性和重复性;将之与基于OIE推荐引物(L2/U2)建立的RT-PCR检测方法进行比较,发现基于引物RP1/RP3检测IBDV的灵敏度明显高于L2/U2,特别是当待检样品中的IBDV的含量低时,会避免因使用L2/U2导致出现漏检的现象。结论:以RP1/RP3作为引物建立的IBDVRT-PCR检测方法灵敏性高、特异性强,可用于IBDV的检疫、诊断和监测。 相似文献
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Newcastle disease virus (NDV), named MET95, was isolated from a non-vaccinated broiler flock in Japan in 1995. The MET95 strain was determined to be a lentogenic NDV. The strain has the properties of eluting rapidly at 4 C and has low thermostability in hemagglutinating activity with chicken erythrocytes. In these studies, no difference could be found between the MET95 strain and the Hitcher B1 vaccine strain. However, the chickens inoculated with the MET95 strain, as well as chickens that they were in contact with, had a much higher hemagglutination-inhibition antibody response than those inoculated with the B1 strain. Accordingly, the MET95 strain is thought to be a promising candidate as a live ND vaccine strain. In Japan, this is the first report on the isolation of lentogenic NDV from chickens since the paper on the Ishii strain isolated in 1966. 相似文献
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In order to explore the influence of nucleoside diphosphate kinase B (NDPK B) on replication and proliferation of F48E9 strain of Newcastle disease virus (NDV), the eukaryotic expression plasmid pEGFP-NDPK B was transfected into Vero cells,then we got the cell subclones by G418 screening. We identified the expression of NDPK B protein by RT-PCR, Western blotting and inverted flurescence microscope. On the basis of Vero/NDPK B cell line,we explored the influence of NDPK B on replication and proliferation of F48E9 strain of NDV by HA-HI, TCID50 and Real-time PCR.The results showed that we had successfully constructed the cell line named Vero/NDPK B, which could stably express NDPK B protein, and found that NDPK B could inhibit the replication and proliferation of F48E9 strain of NDV in Vero cells.It suggested that on the basis of NDPK B,we could design and develop new drugs or antiviral synergist to prevent or treat NDV. 相似文献
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为研究核苷二磷酸激酶B（nucleoside diphosphate kinase B,NDPK B）蛋白表达对新城疫病毒（Newcastle disease virus,NDV） F48E9株复制与增殖的影响,本试验将真核表达重组质粒pEGFP-NDPK B转染Vero细胞,经G418压力筛选出了阳性细胞单克隆,并通过RT-PCR、Western blotting和倒置荧光显微镜观察鉴定了NDPK B mRNA和蛋白质的表达,在细胞系基础上,通过HA-HI、TCID50及实时荧光定量PCR检测NDPK B蛋白对NDV F48E9株复制和增殖的影响。结果表明,试验成功地构建了高表达NDPK B蛋白的Vero/NDPK B细胞系,并在此基础上,通过检测证明了NDPK B能抑制NDV F48E9株在Vero细胞中的复制与增殖,提示以NDPK B为基础有可能设计开发抗NDV的新药物或抗病毒增效剂,对NDV进行预防或治疗。 相似文献
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A rapid, sensitive and specific semi-nested RT-PCR was developed to detect and differentiate virulent and avirulent strains of Newcastle disease virus (NDV). For a total of 67 NDV strains, the results obtained from the semi-nested RT-PCR were consistent with those from nucleotide sequence analysis, plaque forming assays, mean death time (MDT) measurements and intracerebral pathogenicity index (ICPI). Furthermore, 13 class I NDV strains can be characterized by the semi-nested RT-PCR approach, which was feasible by the conventional methods. The detection limit for the semi-nested RT-PCR was two plaque forming units (PFU) both for NDV strain F48E9 in allantoic fluid and for isolate APMV1/ch/ChinaND4031 in oral or cloacal swabs. In conclusion, this semi-nested RT-PCR method offers a new assay for the rapid detection and differentiation of NDVs. 相似文献
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对广东地区疑似发生鸽新城疫感染的鸽病料进行病毒分离,通过血凝试验(HA)、中和试验、F基因扩增及序列测定.结果分离到1株血凝效价为4log2,且能被NDV阳性血清中和的病毒;用针对NDV F基因设计的特异性鉴定引物对该分离株进行PCR扩增,可扩增出相应的目的片段;测序及Blast分析表 明其与山东分离株chicken/... 相似文献
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为了解新疆野鸟新城疫病毒（Newcastle disease virus,NDV）感染情况,分析其分子特征和基因变异特点,防止新城疫的暴发和蔓延,本试验用9日龄SPF鸡胚进行病毒的分离传代,HA、HI试验和RT-PCR方法对位于"东非-西亚迁徙线"福海县的野鸟进行了NDV检测,分离到1株野鸟源NDV,并命名为NDV/Pintail/CH（XJ）/01/2016。结果表明,该NDV分离株F基因ORF长1 662 bp,编码553个氨基酸,F基因碱性裂解位点序列为112G-R-Q-G-R-L117,符合弱毒株特征;遗传进化分析显示,NDV/Pintail/CH（XJ）/01/2016与乌克兰鸽源分离株Doneck/3/968、比利时鸭源分离株Simeonovgrad核苷酸同源性均为99.7%,属于NDV ClassⅡ基因Ⅱ型。而上述3株病毒均与疫苗毒La Sota具有较高同源性（99.5%~99.8%）。本研究结果为家禽NDV外流进入自然环境提供了论据。 相似文献
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ZHANG Hui-min Muhamaitijiang&# T SHI Jun WU Gui-ling Gulizhati&# Adili JIA Er-ken SU Zhan-qiang RAN Duo-liang Shayilan&# Kayizha LAN Ling YAO Gang LIU Jian-hua 《中国畜牧兽医》2017,44(9):2755-2760
In order to understand inflection of Newcastle disease virus (NDV) of Xinjiang wild birds, through analysis of molecular characteristics and genetic variation, to prevent the outbreak and spread of Newcastle disease, isolation of the virus was performed in 9 days old specific pathogen free (SPF) chicken embryos, detected the wild birds in Fuhai county which was located in the migration line of East Africa-West Asia by HA, HI test and RT-PCR method. And then 1 strain of NDV from wild birds was isolated, named as NDV/Pintail/CH(XJ)/01/2016. As the result, ORF of F gene from the NDV isolate was 1 662 bp, encoding 553 amino acids. The motif of the cleavage site was 112G-R-Q-G-R-L117, which was consistent with the characteristics of avirulent NDV strains. The phylogenetic analysis showed that NDV/Pintail/CH(XJ)/01/2016 nucleotide sequence homologies of F gene between reference strain Doneck/3/968 and mule Simeonovgrad strain homology were high and reached 99.7%, it belonged to genotype Ⅱ of Class Ⅱ NDV. All of the three viruses had high homology to the vaccine strain La Sota,which were 99.5% to 99.8%. According to this research conclusion we provided evidence that poultry NDV outflowed into the natural environment. 相似文献