华北地区汉族群体15个短串联重复序列基因座遗传多态性分析

目的调查华北地区汉族人群15个短串联重复序列(shorttandemrepeat,STR)基因座遗传多态性分布和群体遗传学数据。方法应用毛细管电泳技术和五色荧光复合扩增的方法,检测597名汉族无关个体的15个STR基因座基因型。结果15个STR基因座的基因频率分布均符合Hardy-Weinberg平衡,所检测的15个STR遗传标记均具有高度多态性,杂合度均超过0.62,15个基因座的个体识别力在0.802~0.967之间,非父排除率在0.320~0·697之间,匹配概率在0.033~0.198之间。15个基因座的累积个体识别能力为0.999999以上,累积非父排除率为0.99999571,累积匹配概率为8.93×10-18。结论联合检测15个基因座可为亲缘鉴定和个体识别提供可靠的法医学证据,这15个STR基因座适用于中国人群的法医物证学检验。     

四川地区α地中海贫血产前基因诊断

α地中海贫血（α地贫）可分为α地贫1（αα/--）、α地贫2（αα/α-）,缺失型或非缺失型HbH病和Hb Bart′s胎儿水肿综合征（--/--）四种类型。其中以Hb Bart′s胎儿水肿综合征最为严重,可致胎儿缺氧,引起死胎、死产或新生儿死亡,孕妇常有严重的妊娠中毒症及产后出血。因此,及早进行产前诊断,减少此类患儿出生是避免孕妇及其家庭痛苦的有效措施。 除非缺失型HbH病外,α地贫都是由于α基因缺失而致。这种缺失往往引起α基因内限制性内切酶位点的变化,故缺失型α地贫可采用直接检测胎儿DNA上限制性内切酶图谱的方法进行产前诊断。     

用SYBR-Green Ⅰ实时荧光PCR结合融解曲线分析法诊断α-地中海贫血

目的建立自动化、高通量、准确快速检测缺失型α-地中海贫血基因型的技术。方法应用SYBR-Green1进行两个实时荧光聚合酶链反应(real-time fluorescence polymerase chain reaction with SYBR-Green1,SYBR-PCR),检测左缺(-α4.2)、右缺(-α3.7)等位基因,同时进行融解曲线(dissociation curve,DC)和Tm(melting temperature)值分析。PCR产物重组到pCR2.1,重组子梯度稀释作为模板检测灵敏度,确定两种等位基因型(-α4.2,-α3.7)的检测下限,并对110份DNA样品进行检测。结果检测-α4.2和-α3.7的PCR产物长度分别为1.65kb、1.9kb,Tm分别为(81.5±0.5)℃、(82.5±0.5)℃,检测下限分别为9×102个拷贝、4.3×102个拷贝。该检测技术的灵敏度较常规PCR结合琼脂糖凝胶电泳法高10倍。结论SYBR-Green1实时荧光PCR结合融解曲线分析及Tm分析可以灵敏、准确地检测-α4.2、-α3.7、αα(包括αTα)及--SEA4种等位基因,从而为各种缺失型α-地中海贫血做出基因诊断。该技术具有自动化程度高,不需荧光标记探针,成本低,易质控,防污染,高通量等优点,适于临床推广应用。     

Detection of factor Ⅷ intron 1 inversion in severe haemophilia A

Objective Screening the intron 1 inversion of factor Ⅷ (FⅧ) in the population of severe haemophilia A(HA) in China and performing carrier detection and prenatal diagnosis. Methods Using LD-PCR to detect intron 22 inversions and multiple-PCR within two tubes to intron 1 inversions in sereve HA patients. Carrier detection and prenatal diagnosis were performed in affected families. Linkage analysis and DNA sequencing were used to verify these tests. Results One hundred and eighteen patients were seven diagnosed as intron 22 inversions and 7 were intron 1 inversions out of 247 severe HA patients. The prevalence of the intron 1 inversion in Chinese severe haemophilia A patients was 2. 8% (7/247). Six women from family A and 2 from family B were diagnosed as carriers. One fetus from family A was affected fetus. Conclusion Intron 1 inversion could be detected directly by multiple-PCR within two tubes. This method made the strategy more perfective in carrier and prenatal diagnosis of haemophilia A.     

在改进EB作为核酸染色剂中的发现

溴乙锭(EB)是最常见的核酸电泳染色剂,使用该试剂的方法一般有两种,一是将EB加入电泳缓冲液或琼脂糖凝胶中,一是核酸电泳后再在EB溶液中染色。笔者设想电泳前首先将EB与DNA结合为复合物,电泳后就可以直接在紫外灯下观察结果,这不仅背景清晰干净,节约EB用量,减少EB污染,减轻对实验人员的健康危害,也简化了操作。1　材料和方法1.1　溴化乙锭及pBR322/MSPI(0.5μg/μl):亚美公司生产。1.2　方法:1常规铺2%的琼脂糖凝胶。2将溴化乙锭配成10mg/μl的母液,然后分别将母液稀释成1/25,1/50,1/100,各取2μl,与等体积的pBR322/MSPI混合(则…     

酶联免疫吸附法检测肺炎支原体抗体IgG和IgM结果分析

肺炎支原体是人体内支原体属中主要的一种病原体^[1],由其引起的肺炎占X线诊断肺炎的15%～20%^[2],临床上常因诊断不及时而影响治疗,因此对于此种感染的快速诊断是有效治疗的基础。近年发展起来的酶联免疫吸附试验(ELISA)检测肺炎支原体抗体,可同时检测IgG和IgM抗体,特异性强,敏感性高^[3].本研究用ELISA法检测了565例肺炎及上呼吸道感染患者的血清肺炎支原体IgG、IgM抗体。其中的109例同时做冷凝集试验,并对检查结果及治疗情况进行比较分析。     

汉族冠心病人 GPⅢa Pl^A多态性的初步研究

目的：研究血小板膜糖蛋白GPⅢa Pl^A不同基因型在中国汉族人群中的分布情况，探讨Pl^A多态性与汉族人冠心病的遗传易感性的关系。方法：运用多聚酶链反应和限制性内切酶片段长度多态性（PCR—RFLP）技术，对178例冠心病人及130例健康对照组的血小板膜糖蛋白GPⅢa Pl^A进行多态性分析，结合选择性冠状动脉造影结果探讨两者的关系。结果：血小板膜糖蛋白GPⅢa Pl^A等位基因型在冠心病组均表现为Pl^A1/A1纯合型；正常对照组Pl^A的基因型亦表现为Pl^A1/A1型。结论：本组汉族冠心病人血小板膜糖蛋白GPⅢa Pl^A基因型以纯合型Pl^A1/A1型为主；血小板膜糖蛋白GPⅢa Pl^A等位基因型Pl^A2/A2可能不是我国汉族人冠心病的致病因子。     

用基因扩增技术诊断镰状细胞贫血病(简报)

基因扩增,又称PCR(Polymerase Chain Reaction),是近年来国外用于对镰状细胞贫血病及β地中海贫血等已知突变类型的遗传病进行基因诊断及产前基因诊断的新技术。基本方法是:合成可复盖β珠蛋白基因的几对寡核苷酸引物。在每对引物中,一个与编码的DNA链互补,另一个与非编码链互补。每对引物复盖长度为100～300bp。所怀疑的一种或两种点突变位于其中。将1μg人的整基因组DNA在95℃     

α~(32)P-dTTP标记,用末端终止法测定核苷酸排列顺序

用末端终止法测定DNA顺序,并用α-~(32)P-dTTP标记代替α-~(32)P-dATP,摸索了相应试剂系统的合适比例,得到满意的测序效果。     

中国广西,广东,四川区β—地中海贫血基因突变类型及产前基...

beta-Thalassemia is one of the most common single gene disorders in South China, and ten different point mutations and frameshifts have been observed among Chinese. We studied 150 chromosomes of 95 beta-thalassemia patients from Guangxi, Guangdong and Sichuan Provinces using the polymerase chain reaction followed by dot hybridization with specific oligonucleotide probes. The most common mutations were the frameshift at codon 41-42 and the nonsense mutation at codon 17 in Guangxi and Sichuan, and the codon 41-42 frameshift and IVS-II-654 mutation in Guangdong. The A-G mutation at -28 of the promoter was common in Sichuan but not in the other two provinces. Three mutations, -30, IVS-I-1 and IVS-I-5, were not observed. A prenatal diagnosis program using these techniques has been initiated based on these data. Fourteen pregnancies at risk for beta-thalassemia have been diagnosed successfully.