内皮素1基因多态性与原发性高血压的病例对照研究

为探讨重庆市某社区人群内皮素1(Endothelin-1,ET-1)基因G8002A,+138A/-两个位点基因多态性与原发性高血压(Essential hypertension,EH)的相关性,采用PCR-RFLP和PCR-SSP方法分别检测95名EH患者及150名血压正常者组(NT)的G8002A和+138A/-基因多态性.结果发现:在EH组中发现+138A/-位点基因型与舒张压相关(p=0.005),其中4A4A型舒张压比3A3A型高13 mmHg(p=0.005),比3A4A型高12 mmHg(p=0.04).将G8002A与+138A/-位点联合分析后发现舒张压在EH病人中GA4A4A型较其他几型平均高达近20 mmHg,呈显著相关性(p0.01).认为ET-1基因+138A/-的A插入型变异位点可能与重庆市EH人群的舒张压水平有关,GA4A4A基因型人群可能是EH患者血压升高的一个高危遗传因素.     

利用5个多态蛋白（酶）位点分析6个地方猪品种的遗传结构

利用5个多态蛋白（酶）位点碱性磷酸酶（Akp）、蛋白酶抑制物1（PI－1)、后白蛋白1（Po-1）、后白蛋白2（Po-2）和铜蓝蛋白（Cp）分析了清平猪、阳新猪、南阳黑猪、南城黑猪、东乡花猪、杭猪6个地方猪品种的遗传结构和遗传变异。6个品种的平均杂合度大小依次为：南阳黑猪（0.4142）、南城黑猪（0.3057）、阳新猪（0.3032）、东乡花猪（0.2606）、杭猪（0.2478）、清平猪（0.2416），通过计算Nei（1978）标准遗传距离，用UPGMA进行聚类分析，6个品种聚成2大类：清平猪、南城黑猪、阳新猪、南阳黑猪聚成一大类；东乡花猪与杭猪聚成另一类。     

猪钙蛋白酶抑制蛋白基因PCR-RFLP多态性与肉质性状及背膘厚间的关系分析

钙蛋白酶抑制蛋白(CAST)是钙蛋白酶系统的内源性抑制剂，并且已经被确定能影响宰后肉的嫩度。本研究采用PCR-RFLP技术，分析了CAST基因在125头PIC杂交猪中的多态性分布及其与肉质性状和背膘厚间的相关。结果发现，CAST基因在本研究所检测的位点均具有多态性。进一步分析表明，CAST^A2A2基因型个体与其相应的另外两个基因型个体相比肌内脂肪含量有显著差异，CAST^B1B2基因型个体对于肌内脂肪含量的影响也有相同的趋势；CAST^C1C2基因型个体与其相应的另外两个基因型个体相比背膘厚有显著差异，CAST^C1C2。基因型个体和CAST^D1D2基因型个体对于背膘厚的影响也有相同的趋势。因此可以把CAST基因作为影响猪肉盾性状和屠体性状的候选基因。     

水貂14种血液蛋白多态性的研究

采用垂直板聚丙烯酰胺凝胶电泳法对我国饲养的美国短毛黑水貂、大连金州黑水貂和左家型水貂中14个蛋白位点进行检测。结果表明,血液中白蛋白(Alb)、VD结合蛋白(GC)、前白蛋白(Pr)、血红蛋白(Hb)和碳酸酐酶(CA)都是单态,转铁蛋白(Tf)在美国短毛黑水貂和金州黑水貂中呈单态,血清酯酶(Est)由3个共显性等位基因控制,其他位点都有2个控制,后白蛋白(Po)和抗胰蛋白酶(Pi-1)是同一种蛋白。Hardy-Weinberg平衡状态分析表明:Po、抗胰蛋白酶(Pi-3)、慢α球蛋白(Sag)、过氧化物酶(Pox)和(Est)在金州黑水貂中为平衡位点;Po、Pi-3、Sag和Est在左家型水貂中为平衡位点;Po、Pi-3、Pox和Est在美国短毛黑水貂中为平衡位点。平均杂合度的顺序依次为:美国短毛黑水貂>大连金州黑水貂>左家型水貂。3个水貂类型间的亲缘关系,左家型水貂和大连金州黑水貂最近(0.0027),其次是左家型水貂和美国短毛黑水貂(0.0109),金州黑水貂与美国短毛黑水貂最远(0.0213)。这说明利用蛋白多态性识别及探讨水貂品种、品系间的亲缘关系是可行的。     

利用微卫星DNA多态性预测引进肉用绵羊品种杂种优势

【目的】利用微卫星DNA在不同绵羊群体中的多态性预测引进肉用绵羊品种与小尾寒羊的杂种优势。【方法】利用5个微卫星基因座对4个国外引进肉羊品种及小尾寒羊5个群体的等位基因频率、群体多态信息含量、有效等位基因数、杂合度和遗传距离进行了遗传检测，并测定4个引进肉羊与小尾寒的杂交效果。【结果】5个微卫星基因座在白头萨福克、黑头萨福克、道赛特、得克赛儿及小尾寒羊5个群体中存在多态性，可以用于绵羊遗传多样性的评估；从不同品种看，道赛特羊的遗传变异程度最大，而小尾寒羊的遗传变异程度相对较小；经遗传关系分析，在引进的4个肉羊品种中，与小尾寒羊杂交优势由大到小依次为白头萨福克、黑头萨福克、道赛特、得克赛儿，与实际杂种优势测定结果相符。【结论】利用微卫星DNA多态性预测引进肉用绵羊品种与小尾寒羊的杂种优势是可行的，将对今后绵羊育种具有重要的应用价值。     

Acquisition of potato leafroll virus byMyzus persicae from secondarily-infected potato plants of different genotypes

Summary The acquisition of potato leafroll virus (PLRV) byMyzus persicae nymphs from the top leaves of potato plants was studied throughout a growing season in relation to the antigen titre in those leaves and the feeding behaviour of the aphid. Secondarily-infected plants of eight potato genotypes with different levels of field resistance served as virus sources. Early in the growing season, plants were efficient sources for virus acquisition. The amount of viral antigen detected inM. persicae nymphs fed on the top leaves was strongly correlated with the titres of viral antigen in these leaves. Virus acquisition from the top leaves of older potato plants was markedly impaired and could not be correlated with their virus titre. With increasing age of the potato plants and the development of virus symptoms, the virus titre in the leaves declined and the initial weak correlation between the virus titre and field resistance ratings disappeared. Thus, screening secondarily-infected potato plants for field resistance to PLRV based on the concentration of viral antigen in leaves or in aphids fed on them should be avoided later in the growing season. The feeding rate ofM. persicae, measured by the number of honeydew droplets excreted, did not account for the reduced uptake of virus from older plants since it was not influenced by the age of the plant. Throughout the growing season, the feeding rate ofM. persicae nymphs on PLRV-infected plants was higher on genotypes with low levels of field resistance to PLRV than on genotypes with high ones.     

Potato cultivar identification using simple sequence repeats markers (SSR)

Summary Identification of potato cultivars is currently based on phenotypic characters. Crop inspections are needed at different stages and the increasing number of cultivars means the process is becoming more and more complex. Molecular markers are a possible complementary tool to identify potato cultivars and to rapidly check the identity of seeds lots. In this study 286 potato cultivars produced in France were characterized by Polymerase Chain Reaction (PCR) using Simple Sequence Repeats (SSR) markers. Sequential amplifications with 4 to 5 of the chosen SSR markers enabled complete discrimination between all the cultivars. The patterns were registered in a database and the procedure is now used routinely in France.     

非洲猪瘟实时荧光RPA诊断方法建立及应用

【目的】非洲猪瘟（African swine fever,ASF）自2018年首次暴发于我国沈阳后,迅速扩散至全国,对我国养猪业造成了沉重打击。研究针对其病原非洲猪瘟病毒（African swine fever virus,ASFV）建立一种快速核酸检测技术,为及时发现、准确处理ASF疫情提供一种快速诊断方法。【方法】针对ASFV保守的B646L（p72）基因,设计并筛选合适的引物、探针组合,利用基于重组酶-聚合酶扩增（recombinase polymerase amplification,RPA）的实时荧光RPA方法,对反应体系、反应条件、样品处理步骤等进行优化;利用质控品,对检测方法的特异性和灵敏度进行评价。对1 009份临床样品进行检测,并利用OIE推荐的实时荧光定量PCR方法（qPCR）和病毒分离,进一步验证该方法的检测结果。【结果】筛选得到一套合适的引物、探针,建立了针对ASFV p72基因的实时荧光RPA检测方法,优化后的反应体系总体积为25 µL,在实时荧光定量PCR仪器上,最适反应条件为39℃ 10 s,39℃ 20 s,40个循环,扩增反应时间约20 min;室温裂解法可取代传统核酸提取方法作为本检测的样品处理方法;使用优化后的条件,可在30 min内实现样品处理、核酸扩增和结果判定的整个过程,特异性评价结果显示本方法对猪细小病毒（PPV）、伪狂犬病毒（PRV）、圆环病毒1型/2型（PCV1/2）、猪瘟病毒（CSFV）、猪繁殖与呼吸综合征病毒（PRRSV）样品扩增结果均为阴性;敏感性评价结果显示本方法对基因I型、II型、IX型ASFV的模拟样品均可检出,对II型ASFV阳性模拟血样品可检测至10 拷贝/µL,对已知阳性临床组织样品可检至1﹕10^3.0稀释度,与OIE推荐的实时荧光定量PCR（qPCR）方法的检测灵敏度一致。通过对1 009份临床样品进行检测,实时荧光RPA诊断方法检出17份阳性,其结果与qPCR方法完全一致;病毒分离获得17份阳性培养物。【结论】建立的实时荧光RPA核酸检测方法操作简便,耗时短,具有较高的灵敏度和特异性,为临床提供了一种新型、简单、特异、快速诊断非洲猪瘟的方法。     

Establishment of Ecotilling for Discovery of DNA Polymorphisms in Brassica rapa Natural Population

Ecotilling is a new approach based on enzyme-mediated heteroduplex cleavage to discover DNA polymorphisms in natural population. We used mung bean nuclease（MBN） instead of routinely used CELI to cleave single base pair mismatches in heteroduplex DNA templates. Nested set of primers were designed to amplify targeted region to avoid the influence of the variation in quality and quantity of the genomic DNA. To reduce the costs in fluorescently labeled primers, we added M13 adapter to 5＇end of gene specific primers to make IRD dye labeled M13 forward and reverse primers possibly universal for different genes. A Brassica rapa ZIP gene homologue was subjected to the analysis to practise the feasibility of the method in polymorphisms detection. Our experiment showed this method is efficient in discovering DNA polymorphisms in Brassica rapa natural population.     

新疆肠道传播的非甲非乙型肝炎（戊型）病毒（ET—HEV）核酸特异片段的PCR扩增

本研究建立了自患者粪便中直接进行ＥＴ－ＨＥＶ核酸特异片段扩增的方法．即便经反转录反应后的ＰＣＲ放大法。ＰＣＲ流程为：变性：９４℃９０ｓ，退火：４０℃９０ｓ，延伸；７３℃１２０ｓ，共２５个循环。所用引物为ＥＴ－ＨＥＶ特异．从而决定了ＰＣＲ产物的特异性．该产物已用于制备ＥＴ－ＨＥＶＲＮＡ持异的诊断探针．