C-DNA

Abstract: The common species C adenoviruses (serotypes Ad1, Ad2, Ad5, and Ad6) infect more than 80% of the human population early in life. Following primary infection, the virus can establish an asymptomatic persistent infection in which infectious virions are shed in feces for several years. The probable source of persistent virus is mucosa-associated lymphoid tissue, although the molecular details of persistence or latency of adenovirus are currently unknown. In this study, a sensitive real-time PCR assay was developed to quantitate species C adenovirus DNA in human tissues removed for routine tonsillectomy or adenoidectomy. Using this assay, species C DNA was detected in Ficoll-purified lymphocytes from 33 of 42 tissue specimens tested (79%). The levels varied from fewer than 10 to greater than 2 x 10(6) copies of the adenovirus genome/10(7) cells, depending on the donor. DNA from serotypes Ad1, Ad2, and Ad5 was detected, while the rarer serotype Ad6 was not. When analyzed as a function of donor age, the highest levels of adenovirus genomes were found among the youngest donors. Antibody-coated magnetic beads were used to purify lymphocytes into subpopulations and determine whether viral DNA could be enriched within any purified subpopulations. Separation of T cells (CD4/8- expressing and/or CD3-expressing cells) enriched viral DNA in each of nine donors tested. In contrast, B-cell purification (CD19-expressing cells) invariably depleted or eliminated viral DNA. Despite the frequent finding of significant quantities of adenovirus DNA in tonsil and adenoid tissues, infectious virus was rarely present, as measured by coculture with permissive cells. These findings suggest that human mucosal T lymphocytes may harbor species C adenoviruses in a quiescent, perhaps latent form.

Abstract: Publisher Summary This chapter discusses the chemical consequences of radiation damage to DNA Irradiation of DNA causes ionizations in all parts of the DNA molecule, namely, the bases, the sugar-phosphate moiety, and any closely bound water; in addition, bound proteins are also ionized Even though ionizations occur throughout the DNA molecules, initial radicals have been observed only on the DNA bases in low-temperature ESR studies of moist DNA Radiation damage to DNA occurs in a number of ways, which, are classified as either direct or indirect The direct effect corresponds to direct ionization of the DNA resulting in the formation of radical cations and radical anions on the DNA itself The indirect effect corresponds to energy deposition in the surrounding phase followed by attack by radicals from this phase However, in complex systems such as the cell, damage may also be caused by irradiation of molecules bound to the DNA that transfer positive holes and/or electrons to the DNA strands or later react by cross-linking or hydrogen abstraction The quasi-direct effect refers to a third process, where ionizations from irradiation result in holes and dry electrons in biomolecular species and hydration water very near the DNA These then undergo fast transfer to the DNA to form ion radicals on the DNA itself The indirect effect produces many of the same radicals and diamagnetic products as the direct and quasi-direct effects

Abstract: To understand the molecular basis of hemophilia A and to provide heterozygote detection and prenatal diagnosis by DNA analysis, we used cloned factor VIII:C DNA fragments to study 10 affected families. In four of these families, inhibitors of factor VIII:C had developed in affected persons. In one such family a deletion of approximately 80 kb within the factor VIII:C gene was identified. Carriers of the deletion were identified through detection of an abnormal DNA fragment located at the deletion end points. In another family a single nucleotide change in the coding region of the factor VIII:C gene produced a nonsense codon leading to premature termination of factor VIII:C synthesis. Carrier detection was performed in eight female members of this four-generation family. In a third family a small change in the size of a restriction-endonuclease fragment correlated with the presence of the mutant gene, and in the other seven families the molecular defect has not yet been identified. In addition, we used two common polymorphic sites in the factor VIII:C gene to differentiate the normal from the defective gene in four of six obligate female carriers from families with patients in whom inhibitors did not develop. Carrier detection was possible in other members of these families. These data suggest that DNA analysis of the factor VIII:C gene provides an accurate method of carrier detection and, potentially, of prenatal diagnosis in at least 50 per cent of the pedigrees affected by hemophilia A.