A novel type of human papillomavirus associated with genital neoplasias

The role of human papillomaviruses (HPVs) in the development of genital neoplasias has been well documented. The genomes of two HPV types, HPV16 and HPV18, have been found to be associated with about 70% of invasive carcinomas of the uterine cervix. As, under non-stringent hybridization conditions, HPV DNA sequences have been detected in about 90% of cervical carcinomas, it seems likely that additional HPV types are associated with these tumours. Here we report the molecular cloning and characterization of a novel HPV type, tentatively named HPV33, whose sequences have been detected in 4-8% of biopsies of genital intra-epithelial neoplasias and cervical invasive carcinomas, usually as free monomeric or oligomeric molecules. However, in one specimen of vulvar Bowen's disease, HPV33 DNA sequences were integrated in the host-cell genome. Thus, HPV33 probably represents an additional type of potentially oncogenic genital HPV.     

SCGE检测人单个核细胞细胞DNA损伤

用单细胞凝胶电泳法检测了人全血在4℃储存时单个核细胞DNA损伤的情况。结果发现，细胞随储存时间延长基础损伤逐渐增高，储存1，24，48，72h时细胞损伤率分别为8.84%，18.5%，45.2%和54.3%，而细胞对H2O2的敏感性却逐渐降低。以50цmol/L H2O2处理后，其细胞损伤率在0，24，48h分别为63%，61.8%和51.5%。     

用PCR法检测人巨细胞病毒DNA

用聚合酶链式反应（ＰＣＲ）法检测患者尿液中人巨细胞病毒（ＨＣＭＶ）ＤＮＡ．结果表明，自行设计合成的引物位于ＨＣＭＶ基因组早期蛋白基因区，经ＰＣＲ仪扩增一段长４３０ｂｐ的特异序列片段，对正常人基因组ＤＮＡ或其它疱疹病毒ＤＮＡ无交叉反应．此法可检测出少至１０－１６ｇ（０．１ｆｇ）的病毒ＤＮＡ．通过对３５份尿液标本的检测，比较ＰＣＲ法和组织培养法的检测结果完全一致     

DNA化学损伤的电化学检测

0 引言 DNA是重要的遗传物质,在机体的生长过程中,它不可避免地受到各种因素的作用而产生损伤,从而直接影响DNA的复制、转录和蛋白质的合成,进而影响细胞生长、发育、遗传、代谢和繁殖等生命基本过程,还会造成细胞突变、癌变、老化甚至死亡.因此,对DNA损伤及其检测方法的研究不仅有助于更好地了解生命体的基本过程,而且还可为疾病的临床诊断、治疗甚至新的基因治疗药物的发现提供理论基础.     

Morphological transformation in vivo of human uterine cervix with papillomavirus from condylomata acuminata

Carcinoma of the human uterine cervix has been associated with several infectious agents including papillomavirus. Papillomavirus group-specific antigen (GSA) and viral particles have been demonstrated in human condylomata acuminata (CA) and flat warts of the uterine cervix. Cell alterations consisting of nuclear enlargement, hyperchromasia, irregularity, binucleation and cytoplasmic clearing (koilocytosis) are often interpreted as mild to moderate dysplasia. Present evidence that human papillomavirus (HPV) is responsible for the development of these lesions relies on the association of GSA and virus particles in the affected tissue, fulfilling the first two of Koch's postulates. Direct proof of an aetiological relationship, however, requires induction of the CA change in normal, human uterine cervix after exposure to papillomavirus. Infecting human subjects with HPV is ethically unacceptable and no satisfactory alternative systems have been defined. Also, human cell cultures do not support growth or transformation by HPV. Here we report the first demonstration of the morphological transformation of human tissues with a human papillomavirus under controlled, experimental conditions. 'Transformation' is used here in its literal sense to refer to a heritable morphological alteration in the appearance of the cells. The use of this term does not indicate that the changes described are neoplastic, but they are identical to the dysplastic changes found in biopsies of uterine cervical CA. Our results demonstrate the direct involvement of CA virus in dysplastic change of human cervical tissue and indicate that the experimental system described may be useful in elucidating the contribution of human papillomaviruses to the pathogenesis of human cervical cancer.     

金纳米粒子的制备方法及在DNA检测中的应用

综述了金纳米粒子的几种常见制备方法以及这些方法的特点，并归纳为物理法及化学法。前者包括真空蒸镀法、软着陆法及激光消融法；后者又划分为溶胶法、晶种生长法、反胶束法、相转移法及模板法等。不同的方法可获得不同粒径和形状的金纳米粒子，应用时可根据需要选择合适的方法。金纳米粒子特殊的物理及化学性质使其在化学、生物、医学等领域有广泛的用途，尤其在DNA检测中有重要的应用价值。根据方法的特点，DNA检测可分为光学检测法、电化学检测法、压电检测法等。在这些方法中使用金纳米粒子后，检测方法的灵敏度及检测范围有了明显的提高。     

DNA检测方法在鱼类物种鉴定中的应用

总结了近年来在鱼类物种鉴定中常用的DNA序列分析、DNA指纹技术、物种特异性聚合酶链式反应和实时荧光PCR技术等DNA检测方法,同时对各类方法的特点和局限性进行了分析．     

Detection of single base substitutions in total genomic DNA

Certain single base substitutions causing genetic diseases or resulting in polymorphisms linked to mutant alleles, alter a restriction enzyme cleavage site and can therefore be detected in total genomic DNA using DNA blots. Many base substitutions do not lead to an altered restriction site, but these can be detected using synthetic oligonucleotides as hybridization probes if the DNA sequence surrounding the base substitution is known. In the case of beta-thalassaemia, where 22 different single base mutations have been identified, a large number of probes would be required for diagnosis. An approach which was used to detect mutations in viral DNA involves the S1 nuclease treatment of heteroduplexes formed between wild-type and mutant DNA. Although certain single base mismatches are cleaved by S1 nuclease (ref. 11 and T. Shenk, personal communication), many other mismatches examined by this procedure are not cleaved (B. Seed, personal communication; R.M.M., unpublished data). Heteroduplexes between mutant and wild-type subgenomic fragments of double-stranded reovirus RNA migrate slower than the corresponding homoduplexes in polyacrylamide gels containing 7 M urea, but it is not known whether this method is applicable to DNA heteroduplexes containing single base mismatches. Here we describe a procedure that involves the electrophoretic separation of DNA heteroduplexes in a well-characterized gel system. We show that four different human beta-thalassaemia alleles with known single base mutations can be detected with as little as 5 micrograms of total genomic DNA. The method should be useful in the localization and diagnosis of mutations associated with genetic diseases.     

QC-PCR测定HIV-Ⅰ型DNA含量

为人类免疫缺陷病毒（HIV）含量测定提供一种定量的方法。方法 以HIV－Ⅰ型感染小鼠，用竞争性定量多聚酶链式反应（Quantitative Competitive Polymerase Chain Reaction,QC-PCR)和增强化学发光法测定HIV含量。结果 用QC－PCR方法测定HIV含量简单、快速、准确而且无污染。结论 QC－PCR可作为HIV DNA测定的新方法，是AIDS早期诊断、疗效观察及愈后判断的监控手段。     

过氧化草酸酯化学发光检测DNA荧光探针

根据过氧化草酸酯与双氧水作用产生的高能中间体激发荧光物质发光的原理,研究了利用过氧化草酸酯TCPO-H2O2化学发光体系测定Cy5标记的DNA探针的新方法,探讨了溶剂、酸度、催化剂及发光溶液浓度对化学发光信号的影响,确立了最佳分析条件.其线性范围为8.4×10-9~2.8×10-6mol/L,检出限(3σ)为2.8×10-9mol/L.据此建立了操作简便、灵敏度高的测定Cy5标记的DNA探针的新方法.     

Conformational mutations in human mitochondrial DNA

Variation in the human mitochondrial DNA (mtDNA) sequence has been extensively analysed using restriction fragment length polymorphisms (RFLPs). MtDNA RFLPs have previously been attributed to nucleotide changes within restriction endonuclease recognition sites or to small insertion-deletion mutations. We now report that RFLPs detected by polyacrylamide gel electrophoresis can also result from single nucleotide substitutions which alter the mobility of small- to medium-sized restriction fragments that incorporate the sequence. We have defined the mutation responsible at two loci and have identified several possible additional loci. When screening human mtDNAs with multiple restriction endonucleases, such mutations can be misidentified as insertion-deletion mutations or counted as multiple polymorphic restriction sites. This can lead to errors in constructing restriction maps and estimating sequence diversity.     

Hypervariable 'minisatellite' regions in human DNA

The human genome contains many dispersed tandem-repetitive 'minisatellite' regions detected via a shared 10-15-base pair 'core' sequence similar to the generalized recombination signal (chi) of Escherichia coli. Many minisatellites are highly polymorphic due to allelic variation in repeat copy number in the minisatellite. A probe based on a tandem-repeat of the core sequence can detect many highly variable loci simultaneously and can provide an individual-specific DNA 'fingerprint' of general use in human genetic analysis.