Evaluating the outcomes of ethics consultation.
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Consumer Behavior↗
Biomedical subjects
J M Craig
Publications and source records attributed to J M Craig.
Quality of life, justice, and the demands of hospital-based nursing.
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Humans↗
A 330 kb CENP-A binding domain and altered replication timing at a human neocentromere.
Centromere protein A (CENP-A) is an essential centromere-specific histone H3 homologue. Using combined chromatin immunoprecipitation and DNA array analysis, we have defined a 330 kb CENP-A binding domain of a 10q25.3 neocentromere found on the human marker chromosome mardel(10). This domain is situated adjacent to the 80 kb region identified previously as the neocentromere site through lower-resolution immunofluorescence/FISH analysis of metaphase chromosomes. The 330 kb CENP-A binding domain shows a depletion of histone H3, providing evidence for the replacement of histone H3 by CENP-A within centromere-specific nucleosomes. The DNA within this domain has a high AT-content comparable to that of alpha-satellite, a high prevalence of LINEs and tandem repeats, and fewer SINEs and potential genes than the surrounding region. FISH analysis indicates that the normal 10q25.3 genomic region replicates around mid-S phase. Neocentromere formation is accompanied by a replication time lag around but not within the CENP-A binding region, with this lag being significantly more prominent to one side. The availability of fully sequenced genomic markers makes human neocentromeres a powerful model for dissecting the functional domains of complex higher eukaryotic centromeres.
Autoantigens↗
A novel chromatin immunoprecipitation and array (CIA) analysis identifies a 460-kb CENP-A-binding neocentromere DNA.
Centromere protein A (CENP-A) is an essential histone H3-related protein that constitutes the specialized chromatin of an active centromere. It has been suggested that this protein plays a key role in the epigenetic marking and transformation of noncentromeric genomic DNA into functional neocentromeres. Neocentromeres have been identified on more than two-thirds of the human chromosomes, presumably involving different noncentromeric DNA sequences, but it is unclear whether some generalized sequence properties account for these neocentromeric sites. Using a novel method combining chromatin immunoprecipitation and genomic array hybridization, we have identified a 460-kb CENP-A-binding DNA domain of a neocentromere derived from the 20p12 region of an invdup (20p) human marker chromosome. Detailed sequence analysis indicates that this domain contains no centromeric alpha-satellite, classical satellites, or other known pericentric repetitive sequence motifs. Putative gene loci are detected, suggesting that their presence does not preclude neocentromere formation. The sequence is not significantly different from surrounding non-CENP-A-binding DNA in terms of the prevalence of various interspersed repeats and binding sites for DNA-interacting proteins (Topoisomerase II and High-Mobility-Group protein I). Notable variations include a higher AT content similar to that seen in human alpha-satellite DNA and a reduced prevalence of long terminal repeats (LTRs), short interspersed repeats (SINEs), and Alus. The significance of these features in neocentromerization is discussed.
Alu Elements↗
Quantitative microscopy after fluorescence in situ hybridization - a comparison between repeat-depleted and non-depleted DNA probes.
Complex probes used in fluorescence in situ hybridization (FISH) usually contain repetitive DNA sequences. For chromosome painting, in situ suppression of these repetitive DNA sequences has been well established. Standard painting protocols require large amounts of an unlabeled 'blocking agent', for instance Cot-1 DNA. Recently, it has become possible to remove repetitive DNA sequences from library probes by means of magnetic purification and affinity PCR. Such a 'repeat depleted library probe' was hybridized to the q-arm of chromosome 15 of human metaphase spreads and interphase cell nuclei without any preannealing by Cot-1 DNA. Apart from this, 'standard' FISH conditions were used. After in situ hybridization, microscope images were obtained comparable to those achieved with the #15q library probe prior to depletion. The images were recorded by a true color CCD camera. By digital image analysis using 'line scan' and 'area scan' procedures, the painting efficiency expressed in terms of relative fluorescence signal intensity was quantitatively evaluated. The painting efficiency using the repeat depleted probe of chromosome 15q was compared to the painting efficiency after standard FISH. The results indicate that both types of probes are compatible to a high FISH efficiency. Using equivalent probe concentrations, no significant differences were found for FISH with standard painting probes and repeat depleted painting probes.
Cell Nucleus↗
Seizure possibly associated with fluvoxamine.
OBJECTIVE: To inform clinicians of the possibility that seizures due to therapeutic doses of fluvoxamine may not be as rare as previously considered. CASE SUMMARY: A 49-year-old white man with schizoaffective disorder and a past history of seizures secondary to head trauma had been seizure-free for approximately 10 years. Fluvoxamine therapy was begun due to increasing obsessive-compulsive behavior. Despite receiving anticonvulsants for his mood disorder, the patient had a breakthrough seizure. There were no underlying medical conditions that might have induced this seizure. No further seizures occurred after he was placed on a higher dosage of the anticonvulsants. The obsessive-compulsive behavior improved considerably as a result of fluvoxamine treatment. DISCUSSION: The patient presented here developed a seizure with a therapeutic dosage of fluvoxamine; seizures associated with this agent have occurred more often with overdose. Multiple factors such as a prior history of seizures, head trauma, and concurrent treatment with other psychotropic agents are considered in this case report. CONCLUSIONS: Despite the relatively safe and benign adverse effect profile of the selective serotonin-reuptake inhibitors such as fluvoxamine, clinicians should be cautious about seizures as an adverse effect, especially when the patient has even a remote history of seizure or head trauma.
Dose-Response Relationship, Drug↗
Mammalian centromeres: DNA sequence, protein composition, and role in cell cycle progression.
The centromere is a specialized region of the eukaryotic chromosome that is responsible for directing chromosome movements in mitosis and for coordinating the progression of mitotic events at the crucial transition between metaphase and anaphase. In this review, we will focus on recent advances in the understanding of centromere composition at the protein and DNA level and of the role of centromeres in sister-chromatid cohesion and mitotic checkpoint control.
Animals↗
A complete set of repeat-depleted, PCR-amplifiable, human chromosome-specific painting probes.
We describe the generation of a complete set of human chromosome-specific painting probes depleted in repetitive sequences. These probes yield highly specific signals when hybridized without the addition of a blocking agent, such as Cot-1 DNA, and without probe preannealing prior to hybridization. Fluorescent intensities and signal-to-background ratios for these probes are comparable to those of untreated probes hybridized with Cot-1 DNA. We demonstrate the suitability of these probes for applications with very complex probe sets, such as multiplex-FISH.
Binding, Competitive↗
Word recognition performance for Northwestern University Auditory Test No. 6 word lists in quiet and in competing message.
Word recognition norms were determined for the Northwestern University Auditory Test No. 6 (NU-6) released on the Department of Veterans Affairs (VA) Disc 1.1. Word recognition performance (in quiet and in competing message) was measured at 12 presentation levels for 24 young adults with normal hearing. Test-retest reliability also was evaluated. Word recognition scores for VA Disc 1.1 were generally higher than those reported for VA Disc 1.0. The differences in NU-6 scores across disc versions, however, were small and unlikely to affect clinical decisions based on word recognition tests. Score differences on test-retest for the VA Disc 1.1 version of the NU-6 with a competing message background also were small and unlikely to affect clinical outcomes. Overall, based on comparisons of scores for the disc versions of the NU-6, it appears that the two different recordings can be used interchangeably for clinical applications.
Adolescent↗
The chromosomal distribution of CpG islands in the mouse: evidence for genome scrambling in the rodent lineage.
We have prepared a library of mouse whole CpG islands using a methyl-CpG binding domain column. The distribution of CpG islands in the mouse genome was determined by FISH, using the library as a probe. Unlike in other vertebrate genomes that have been examined (human and chicken), extreme clustering of CpG islands was not seen in the mouse genome. No individual murine chromosome stood out as being either very gene-rich or very gene-poor. Despite the more even distribution of CpG islands in the mouse at a gross chromosomal level, at finer resolution concentrations of CpG islands are seen to correspond to the R-band early replicating regions of the genome.
Animals↗
Removal of repetitive sequences from FISH probes using PCR-assisted affinity chromatography.
The vast majority of probes used in fluorescence in situ hybridization (FISH) contain repetitive DNA. This DNA is usually competed out of a hybridization reaction by the addition of an unlabeled blocking agent, Cot-1 DNA. We have successfully removed repetitive DNA from two complex FISH probe sets: a degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR) single human chromosome library and genomic DNA. The procedure involved hybridizing in solution a DOP-PCR-amplifiable probe set with a 50-fold excess of biotin-labeled Cot-1 DNA, and capturing the Cot-1 DNA-containing hybrids using streptavidin magnetic particles, followed by purification and reamplification of the unbound fraction. Probes were checked for depletion of repeats by hybridization to chromosomes without Cot-1 DNA. Results showed hybridization patterns comparable to those achieved with untreated probes hybridized with Cot-1 DNA.
Chromatography, Affinity↗
Semen donors and STD screening.
AIM: The British Andrology Society recommends screening semen donors for sexually transmitted infections to minimise the risk of pathogen transmission to the mother and fetus. The aim was to review recent findings of semen donor screening and, if appropriate, recommend changes to the screening protocol. SUBJECTS: 175 consecutive men attending for STD screening between January 1992 and December 1995 who had been preselected by the Department of Obstetrics and Gynaecology as suitable semen donors. METHODS: Retrospective review of case notes and group comparison of demographic and sexual history data. RESULTS: 11 men (6%) had evidence of infection, excluding CMV seropositivity, at their first STD screen. After semen donation, 109 men (63%) were rescreened and, of these, 12% had positive findings. Positive findings at initial screening were predicted by a history of more than one partner in the preceding 6 months (OR 7.11, 95% CI 1.66-30.4) but it did not predict rescreening findings. Other factors such as age, marital status, employment status or past STDs were not predictive for either screen. DISCUSSION: Less than 20% of initial volunteers meet the full criteria of high quality post-thaw semen, no transmissible genetic disorders, and no transmissible pathogens. Sexual history may predict but would not alone preclude all positive STD screening findings. It is essential that sequential STD screening of donors continues and that genitourinary physicians should be involved in this process. Validation of newer diagnostic techniques as screening tests in this setting is required.
Adult↗
Azole resistance in yeasts.
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AIDS-Related Opportunistic Infections↗
Scaffold attachments within the human genome.
It is generally agreed that, above the level of the 30 nm fibre, eukaryotic chromatin is constrained into loops, but there is disagreement about the nature of the substructure that serves to anchor loops and the DNA sequences that act as the attachment sites. This problem may stem from the very different methods that all purport to separate loop and attached DNAs. We have tested ideas about how the genome is arranged into loops by analysing the average loop size over different cytologically resolvable regions of human chromosomes using fluorescence in situ hybridisation with loop and attached DNA fractions. Variations in average loop size, along and between chromosomes, measurable at this level of resolution were small but significant and were dependent on the extraction method. This emphasises the fundamental differences between the nuclear substructure probed by different protocols. DNA attached to the nuclear 'scaffold' or 'matrix' hybridises preferentially to gene-poor regions of the genome (G-bands). Conversely, fractions attached to the nuclear 'skeleton' hybridise preferentially to gene-rich R-bands and sites of high levels of transcription. The inactive X chromosome has a deficit of associations with the nuclear skeleton but not with the matrix or scaffold. A large excess of attached sequences is found at some sites of constitutive heterochromatin, but not at centromeres.
Cell Nucleus↗
Factors associated with elevated cutaneous Malassezia pachydermatis populations in dogs with pruritic skin disease.
The prevalences of breeds and concurrent diseases in a group of 40 dogs with pruritic skin disease associated with elevated cutaneous Malassezia pachydermatis populations were compared with samples of a dermatological hospital population. The ages and genders of the affected dogs were comparable to those of the dermatology population. Basset hounds, cocker spaniels and West Highland white terriers were significantly overrepresented. Concurrent diseases were diagnosed in 27 dogs, of which 15 were atopic. However, the prevalences of atopic disease, primary keratinisation defects and endocrinopathies in dogs with elevated cutaneous M pachydermatis populations were comparable to those in the dermatology population as a whole. These results indicate that certain breeds are predisposed to the development of elevated cutaneous M pachydermatis populations and that concurrent skin diseases can frequently be identified in affected dogs. However, the relationship between these concurrent diseases and abnormal M pachydermatis populations remains unclear.
Animals↗
The distribution of CpG islands in mammalian chromosomes.
Using fluorescent in situ suppression hybridization to metaphase chromosomes, we have directly shown that CpG islands are predominantly found in the early replicating (R band) regions of the genome. Conversely, late replicating (G band) DNA is sparsely populated with islands. The very highest concentration of CpG islands is in a subset of R bands, most of which are known as T bands. We suggest that there is an interdependence between the differences in island density and the behaviour of chromosomal domains. Our findings indicate which regions of the genome will yield the highest density of coding sequence information. An awareness of local island density may influence the choice of method for identifying exons in genomic DNA.
Animals↗
Chromosome bands--flavours to savour.
The mammalian chromosome is longitudinally heterogeneous in structure and function and this is the basis for the specific banding patterns produced by various chromosome staining techniques. The two most frequently used techniques are G, or Giemsa banding and R, or reverse banding. Each type of stained band is characterised by variations in gene density, time of replication, base composition, density of repeat sequences, and chromatin packaging. It is increasingly apparent that R and G bands, which are complementary to each other, represent separate compartments of the euchromatic human genome, with R bands containing the vast majority of genes. R bands are also more GC-rich, contain a higher density of Alu repeats, and replicate earlier in S phase, than G bands. These properties may be interdependent and may have coevolved.
Base Composition↗