人酪氨酸羟化酶基因真核表达载体的构建及在哺乳动物细胞中的表达

用限制性内切酶EcoRI从pKS(-)HTH_1切下全长为1.9 kb的人酪氨酸羟化酶基因,在T_4DNA连接酶的作用下连接在真核表达载体pCDNA_3的EcoR Ⅰ位点,构建成重组质粒pcD-NA_3HTH_1,该质粒转染COS-7细胞,免疫荧光组织化学染色证实酪氨酸羟化酶在其中的表达。     

The Construction fo Hybrid Plasmid pBR322:RSF1010

The plasmids pBR322 and RSF1010,extracted from E.coli HB802 and E.coliC600 respectively,were treated with restrictive endonuclease EcoRI andligated with T4 DNA Iigase.C600 strains were then transformed with thehybrid plasmid(pSMMl),and thus253transformants were obtained.Thefigures of gel electrophoresis and electromicrograph proved that pSMMlwas a pBR322:RSF1010 composite hybrid.In E.coli C600 pSMMl obtainedresistance against ampieilline(Ap),tetracycline(Tc)and streptomycin(Sm).The orientation of pBR322 and RSF1010 in hybrid pSMMl and the failureof introduction of this hybrid into P.putida AC10 were discussed.     

抗氧化性维生素与男性生殖

生殖系统内过多的活性氧 (ROS)产生可导致精子膜脂质过氧化损伤 ,而抗氧化性维生素对于清除ROS 避免精子脂质过氧化损伤具有一定保护作用。本文就抗氧化性维生素在男性生殖中的作用及其在男性不育治疗等方面进行了综述     

男性不育患者精浆尿酸的检测及临床意义初探

目的 :检测男性不育患者精浆尿酸的含量 ,并探讨其与不育的关系。　方法 :2 0 0 3年 2～ 8月就诊的男性不育患者 1 6 3例 ,分为 4组 :梗阻性无精子症组 ,1 5例 ;非梗阻性无精子症组 ,36例 ;少精子症组 ,4 3例 ;弱精子症组 ,6 9例。 2 0例正常生育男性为正常对照组。上述各组均作精液参数分析及精浆尿酸含量的测定。　结果 :正常对照组精浆尿酸含量为 (396 .9± 5 3.1 ) μmol/L ,显著高于梗阻性无精子症组 [(79.5± 1 8.1 ) μmol/L]、非梗阻性无精子症组[(2 4 5 .8± 76 .5 ) μmol/L]、少精子症组 [(2 6 2 .2± 79.2 ) μmol/L]和弱精子症组 [(2 5 1 .4± 75 .4 ) μmol/L](P均 <0 .0 1 )。其中 ,梗阻性无精子症组精浆尿酸含量又显著低于其他各不育症组 (P均 <0 .0 1 ) ,其余各不育症组间精浆尿酸含量差异无显著性 (P >0 .0 5 )。　结论 :精浆中尿酸作为生殖系统中的一种重要抗氧化物 ,可能在男性生殖中具有一定意义。     

Results of a high-resolution genome screen of 437 Alzheimer's disease families

Alzheimer's disease (AD) is a devastating neurodegenerative disorder of late life with complex inheritance. Mutations in three known genes lead to the rare early-onset autosomal dominant form of AD, while a common polymorphism (epsilon 4) in the gene encoding apolipoprotein E (APOE ) is a risk factor for more typical late-onset (>60 years) AD. A recent study concluded that there are up to four additional genes with an equal or greater contribution to the disease. We performed a 9 cM genome screen of 437 families with AD, the full National Institute of Mental Health (NIMH) sample, which has been carefully ascertained, evaluated and followed by our group over the last decade. Performing standard parametric and non-parametric linkage analyses, we observed a 'highly significant' linkage peak by Lander and Kruglyak criteria on chromosome 19q13, which probably represents APOE. Twelve additional locations-on 1q23, 3p26, 4q32, 5p14, 6p21, 6q27, 9q22, 10q24, 11q25, 14q22, 15q26 and 21q22-met criteria for 'suggestive' linkage [i.e. two-point lod score (TLS) >/=1.9 and/or multipoint lod score (MLS) >/=2.2] in at least one of our analyses. Although some of these will surely prove to be false positives, these linkage signals should provide a valuable framework for future studies aimed at identifying additional susceptibility genes for late-onset AD.     

Identification of a type 1 diabetes-associated CD4 promoter haplotype with high constitutive activity

CD4 is a candidate gene in autoimmune diseases, including Type 1 diabetes mellitus (T1DM), because the CD4 receptor is crucial for appropriate antigen responses of CD4(+) T cells. We previously found linkage between a CD4-1188(TTTTC)(5-14) promoter polymorphism and T1DM. In the present study, we screened the human CD4 promoter for mutations and identified three frequent single nucleotide polymorphisms (SNPs): CD4-181C/G, CD4-521C/G and CD4-1050T/C. The SNPs are in strong linkage disequilibrium (LD) and association with the CD4-1188(TTTTC)(5-14) alleles, and we observed nine CD4 promoter haplotypes, of which four are frequent. We genotyped the SNPs in 253 Danish T1DM families (1129 individuals) and found evidence for linkage and association of a CD4 (A4(-1188)T(-1050)G(-521)C(-181)) haplotype to T1DM. In reporter studies, we show that (1) the T1DM-associated CD4 haplotype encodes high constitutive promoter activity and (2) the CD4-181G variant encodes higher stimulated promoter activity than the CD4-181C variant. This difference is in part neutralized in the frequently occurring CD4 promoter haplotypes by the more upstream genetic variants. Thus, we report functional impact of a novel CD4-181C/G SNP on stimulated CD4 promoter activity and the identification of a novel CD4 haplotype with high constitutive promoter activity that is linked and associated with T1DM.     

No evidence for transmission disequilibrium between a new marker at the myelin basic protein locus and multiple sclerosis in French patients

The myelin basic protein (MBP) gene is a candidate locus for susceptibility to multiple sclerosis. Several groups have tested a complex (TGGA)n repeat in the 5' region of this gene for association/linkage with multiple sclerosis, with divergent results. This region of tandem repetitive sequence has been subjected to complex rearrangements, and there is a possibility that alleles of the same size have different internal structures, which reduces the interest of this marker for linkage disequilibrium studies and may at least partly explain the conflicting results obtained so far. To overcome this problem, we isolated a new polymorphic (CA)n repeat within the Golli-MBP locus. The limited number of alleles identified makes this other marker suitable for transmission disequilibrium studies. We tested this marker for linkage with multiple sclerosis, using the transmission-disequilibrium test (TDT) on a sample of 196 nuclear families in which the genotypes of both parents could be unambiguously defined. We found no evidence of transmission disequilibrium between multiple sclerosis and any of the three alleles of this marker, even when the patients were subdivided according to their HLA-DRB1*1501 status. The present data thus provide no evidence for a contribution of the MBP gene to multiple sclerosis susceptibility in French patients.     

亚甲基四氢叶酸还原酶基因多态性与冠心病关系的研究

目的探讨Ｎ５－亚甲基四氢叶酸还原酶（ＭＴＨＦＲ）基因多态性与冠状动脉粥样硬化性心脏病的关系。方法利用聚合酶链反应和限制性片段长度多态性（ＰＣＲ－ＲＦＬＰ）方法，检测了７９名健康人和６９名冠心病患者ＭＴＨＦＲ基因的６７７碱基多态性突变Ｃ→Ｔ情况，并加以对照分析。结果冠心病患者ＭＴＨＦＲ基因突变型Ｖ６７７基因的频率，明显高于健康人，差异极显著（Ｐ＜０．０１）。结论提示ＭＴＨＦＲ基因突变型Ｖ６７７基因可能是冠心病的又一个遗传风险因子，为探讨冠心病发病机理提供了新的依据     

遗传代谢病所致贫血的诊疗专家共识

遗传代谢病是一大类复杂的单基因病,由于代谢通路异常导致物质代谢紊乱,可以引起单系统或多系统损害.血液系统疾病涉及红细胞、白细胞和血小板功能和数量的异常,其中贫血最常见,新生儿到老年均可发病.现有研究发现,多种遗传代谢病可导致贫血,致病机制不同,一些疾病引起铁、维生素B12、叶酸代谢异常,另一些造成红细胞膜、代谢相关酶或血红蛋白合成异常,引起不同类型不同程度的贫血.为了提高临床医生对遗传代谢病所致贫血及血液系统疾病的认识,早期诊断、精准治疗、改善预后,国内多学科专家共同讨论,结合国内外经验及指南,以巨幼细胞性贫血、溶血性贫血、铁粒幼细胞性贫血和小细胞低色素性贫血四类贫血相关的遗传代谢病的病因、发病机制、临床表现和治疗为重点,制定了本共识,以期指导优化临床诊治实践.     

Effects of CCR5-Delta32 and CCR2-64I alleles on HIV-1 disease progression: the protection varies with duration of infection

OBJECTIVE: To examine temporal variation in the effects of CCR5-Delta32 and CCR2-64I chemokine receptor gene polymorphisms on HIV-1 disease progression. DESIGN: Pooled analysis of individual patient data from 10 cohorts of HIV-1 seroconverters from the United States, Europe, and Australia. METHODS: We studied HIV-1 seroconverters of European (n = 1635) or African (n = 215) ancestry who had been genotyped for CCR5-Delta32 and CCR2-64I. We used Cox proportional hazards models with time-varying coefficients to determine whether the genetic protection against AIDS (1987 case definition) and death varied with time since seroconversion. RESULTS: Protection against AIDS conferred by CCR5-Delta32 held constant at a 31% (RH 0.69, 95% CI 0.54, 0.88) reduction in risk over the course of HIV-1 infection, whereas protection against death held constant at a 39% reduction in risk (RH 0.61, 95% CI 0.45, 0.88). When the period from AIDS to death was isolated, the survival benefit of CCR5-Delta32 diminished 2 years after AIDS. Protection against AIDS conferred by CCR2-64I was greatest early in the disease course. Compared with individuals without CCR5-Delta32 or CCR2-64I, individuals with one or two copies of CCR2-64I had a 58% lower risk of AIDS during the first 4 years after seroconversion (RH 0.42, 95% CI 0.23, 0.76), a 19% lower risk during the subsequent 4 years (RH 0.81, 95% CI 0.59, 1.12), and no significant protection thereafter. CONCLUSION: The protection against AIDS provided by CCR5-Delta32 is continuous during the course of infection. In contrast, the protection provided by CCR2-64I is greatest early in the course of infection.