重庆市山羊博尔纳病病毒P24基因的检测

为了解重庆市山羊博尔纳病隐性带毒情况,分析博尔纳病病毒(BDV)的种系来源。采用巢式逆转录酶PCR结合荧光定量PCR(FQ-nRT-PCR)技术对重庆市60只山羊外周血单核细胞 (PBMCs)及脑组织中的BDV P24基因片段进行了检测,将阳性产物测序,并与国外BDV毒株进行了比较。结果,山羊外周血检测阳性率为8．3％(5／60),脑组织检测阳性率为10％(6／60)。该 BDV P24片段核苷酸序列与马源BDV H1766株同源性最高,达96．51％,与标准株Strain V和 He／80同源性为95．35％,并且编码的氨基酸序列也相同。表明,重庆市山羊中存在动物源性博尔纳病隐性带毒。该BDV P24核苷酸序列与Strain V和He／80株具有高度同源性。     

丙酸钠和丙酮酸钠对体外培养牛肝细胞内丙酮酸羟化酶基因mRNA水平的影响

在成功构建牛肝细胞培养模型和PC基因DNA竞争模板的基础上,采用竞争RT-PCR方法检测了丙酸钠和丙酮酸钠对体外培养新生牛单层肝细胞PC基因mRNA丰度的影响。结果,随着丙酸钠浓度的升高,PC基因mRNA水平呈上升趋势,PC基因mRNA对丙酸钠的耐受范围较广;随着丙酮酸钠浓度的升高PC基因mRNA水平呈先上升后下降的趋势。结果表明,肝细胞内PC基因mRNA的表达水平受丙酮酸钠浓度的调控,在一定范围内丙酮酸钠对PC基因mRNA的转录具有促进作用,而浓度过高时则起抑制作用。     

利用转基因植物生产口蹄疫可饲疫苗的研究进展与前景

从口蹄疫可饲疫苗的研究概况、生产技术要点、优缺点及需要改进的问题几方面阐述了利用植物反应器生产口蹄疫可饲疫苗的研究前景。综述了口蹄疫可饲疫苗的研究进展,介绍了利用植物反应器生产口蹄疫可饲疫苗的技术要点,包括在口蹄疫病毒遗传转化质粒载体系统中最常用的农杆菌介导法,提出了生产口蹄疫可饲疫苗需要改进的技术问题。     

H9N2亚型禽流感病毒聚合酶基因的克隆及序列分析

采用RT-PCR方法对1株H9N2亚型禽流感病毒Ck/GS/2/99株RNA聚合酶基因PB2、PB1和PA分别进行了扩增,将其克隆到pGEM-T easy载体后进行序列测定与分析。结果表明,该株病毒的PB2、PB1和PA基因开放阅读框(ORF)分别由2280、2274和2 151个碱基组成,分别编码759、757和716个氨基酸。经同源性与系统发生树分析,该毒株RNA聚合酶基因PB2、PB1和PA与Ck/NX/4/99株的核苷酸和氨基酸同源性均在98%以上,另外这3个基因与Dk/HK/Y280/97、Sw/HK/9/98和Qu/HK/NT/28/99株的关系密切。     

扩增X—Y同源Amelogenin基因内含子在性别鉴定中的应用研究

本文用一对针对X－Y同源的Amelogenin基因第一内含子的引物，于同一试管中分别扩增出针对于x和Y染色体的特异性DNA片段：106bp及112bp的PCR产物，经聚丙烯酰胺凝胶电泳后用银染法观察扩增结果，取得了满意的效果．对50pgDNA模板室温放置16年的血痕及单根毛发、烟头等检材均可得到明确的性别鉴定结果．本实验证明该方法简便、灵敏、可靠、特别适用于腐败降解检材的法医学性别鉴定．     

An innovative transfer DNA experimental design and qPCR assay to identify primary,secondary, and tertiary DNA transfer

“Touch DNA” is a form of trace DNA that is presumed to be deposited when an individual touches something and leaves behind DNA-containing skin cells, sweat, or other fluids. While touch DNA is often the result of direct contact (i.e., primary transfer), it can also be indirectly transferred between surfaces or individuals (e.g., secondary or tertiary transfer). Even experts cannot distinguish between different types of transfer and do not fully understand which variables affect direct versus indirect transfer or how often each type of transfer occurs. In this study, we utilize an innovative protocol that combines a paired male and female transfer DNA experimental design with an Amelogenin qPCR assay to generate data on primary, secondary, and tertiary DNA transfer. We report frequencies of indirect DNA transfer and also investigate the potential effects of participant age, self-identified ethnicity, and skin conditions on DNA transfer. Out of 22 experimental trials, we detected primary transfer (male + female) in 71% of trials, secondary DNA transfer in 50% of trials, and tertiary DNA transfer in 27% of trials. No significant associations were found between primary DNA transfer and age, self-identified ancestry, or skin conditions, however, all individuals with sloughing skin conditions demonstrated primary DNA transfer and we suggest this variable be explored in larger samples. These results contribute to a better understanding of the conditions under which secondary and tertiary DNA transfer occurs and can be used to propose realistic DNA transfer scenarios in court cases.     

Death Due to Complications of Anhidrotic Ectodermal Dysplasia

Ectodermal dysplasia comprises a group of disorders affecting ectodermal tissues. Severity depends on the genetic aberration; hyperpyrexia secondary to absence of sweat glands is a common complication. Treatment is supportive. This case report describes a 1‐month, 27‐day‐old male infant with a diagnosis of X‐linked recessive anhidrotic ectodermal dysplasia. On the day of his death, his mother swaddled him in a blanket and placed him on the couch at 5:30 am. When she picked him up at 8:00 am, he was unresponsive. At the emergency department, his rectal temperature was 40°C. Postmortem blood culture was positive for group B streptococcus, a possible etiology for fever. It is vital to teach parents that close monitoring of children with ectodermal dysplasia is necessary, as an increase in body temperature can become life threatening.     

非洲猪瘟TaqMan实时荧光定量PCR检测方法的建立与应用

为建立检测非洲猪瘟病毒(African swine fever virus,ASFV)的快速诊断方法,本研究根据ASFV SY18株p17蛋白的编码基因D117L的保守序列设计并合成引物和探针,建立了基于ASFV p17蛋白的编码基因D117L的TaqMan荧光定量PCR检测方法,并验证其特异性、灵敏性和重复性。结果显示,本研究建立的TaqMan荧光定量PCR检测方法的C_t值与标准品在1×10^9~1×10^1copies/μL范围内呈良好的线性关系,相关系数为0.998,斜率为-3.192,检测下限为10 copies/μL,且与其他能引起相似症状的猪源病毒无交叉反应。重复性试验结果显示,组间与组内变异系数均小于1.920 7%,重复性好。此方法可用于ASF的早期诊断和ASFV快速检测。     

PowerPlex~ 21试剂盒扩增后的分型图谱中Y片段丢失的识别

目的依据分型图谱,直观、简便识别是否存在Amelogenin基因Y片段丢失的可能。方法采用计算分型图谱中等位基因峰高之和比值的方法,统计1 968份人员样本经Power Plex~ 21试剂盒扩增后Amelogenin与D3S1358基因座之间的均衡性、Amelogenin X等位基因与Amelogenin Y等位基因的均衡性以及D3S1358基因座不同等位基因的均衡性情况。结果 90.8%的女性样本Amelogenin X等位基因峰高不低于D3S1358基因座等位基因峰高和的60%,94.9%的男性样本Amelogenin X等位基因的峰高不超过D3S1358基因座等位基因峰高和的70%。结论 Power Plex~ 21试剂盒扩增后的分型结果中,当Amelogenin基因只检测到Amelogenin X等位基因,且Amelogenin X等位基因的峰高不及D3S1358基因座等位基因峰高之和的70%时,应考虑存在Y片段丢失的可能性。     

大鼠急性肾缺血再灌注损伤早期的基因表达

目的研究急性肾缺血再灌注损伤早期的差异基因表达,探索其潜在的分子机制。方法通过单纯夹闭大鼠肾动脉制备肾缺血再灌注模型,进行基因表达芯片检测和生物信息学分析,筛选急性肾损伤早期的差异基因表达及其所涉及的细胞活动和信号通路。同时,通过构建差异表达基因的生物网络来寻找与急性肾损伤关系密切的分子,并进行荧光定量PCR验证。结果在检测出的151个差异表达基因中,132个基因显著上调,19个基因显著下调,GO与KEGG通路富集分析结果显示主要与细胞增殖、细胞因子介导的信号通路和免疫反应等有关。荧光定量PCR结果显示,与对照组相比,挑选出的3个与急性肾损伤高度相关的基因(ANXA1、PHLDA1、KLF6)在疾病早期具有显著的表达差异,上升倍数与芯片检测出的倍数基本一致。结论本研究揭示了急性肾损伤早期的差异基因表达特征以及所涉及的生物学过程和信号通路,其中ANXA1、PHLDA1、KLF6可能在急性肾损伤的发病机制中起到一定作用。