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R K Wilson

Publications and source records attributed to R K Wilson.

16 recordsLinked to original sources

Fluorescence chemistries for automated primer-directed DNA sequencing.

A major limitation in the applicability of automated DNA sequencing instruments has been the difficulty in using user-defined oligonucleotide primers which allow sequencing reactions to start at any specific point in a region of interest. Recently, new chemistries have become available for fluorescent labeling which will begin to facilitate the use of any oligonucleotide primer with automated DNA sequencers. In this report, we describe several methods for automated primer-directed DNA sequencing, and compare and discuss the relative merits and limitations of these methods.

Automation

Nucleotide sequence analysis of 95 kb near the 3' end of the murine T-cell receptor alpha/delta chain locus: strategy and methodology.

The nucleotide sequence of a region at the 3' terminus of the murine T-cell receptor alpha/delta chain locus is presented. This region, which encodes the constant region genes for alpha and delta chain polypeptides and all 50 joining gene segments for the alpha chain polypeptide, spans 94,647 bp and includes more than 50 noncoding sequence elements important for T-cell receptor gene rearrangement and expression. DNA sequencing of this region included complete analysis of two cosmid clones and five additional restriction fragments using a random subcloning approach with various manual and automated sequencing strategies. The automated sequencing strategies hold considerable promise for future large-scale DNA sequencing efforts.

Animals

Organization, structure, and function of 95 kb of DNA spanning the murine T-cell receptor C alpha/C delta region.

We have analyzed the organization, structure, and function of the murine T-cell receptor C alpha/C delta region. This region spans 94.6 kb of DNA and contains the C alpha and C delta genes, as well as the V delta 5, J delta 2, and 50 different J alpha gene segments. Within this sequence we have identified 15 new J alpha gene segments, 40 new 5' RNA splice signals, and 40 new DNA rearrangement signals for the J alpha gene segments. The murine C alpha/C delta sequence contains an exceptionally high level of coding sequence with over 5.7% of the total sequence found in the exons. This is much more than that found in the beta-globin locus and the HPRT locus. Using the sequence data obtained from the C alpha/C delta region, we have designed simple assays to test for J alpha gene segment transcription and to determine the level of polymorphism for simple repeat sequences among different inbred strains of mice using the polymerase chain reaction. Furthermore, comparisons of this 95 kb of sequence with the available sequence from homologous regions of other species have led to the identification of a highly conserved sequence that is present throughout vertebrates and in the mouse binds lymphocyte-specific nuclear proteins. Comparisons of a 10-kb region, which includes the C alpha gene in human and mouse, average 66% sequence similarity. These studies support the contention that large-scale DNA sequencing projects of homologous regions of mouse and human will provide powerful new tools for studying the biology and evolution of loci such as the T-cell receptor and for identifying and posing new questions about the functions of conserved sequences.

Amino Acid Sequence

Amantadine aerosol in humans.

Seven well volunteers and three patients with a naturally occurring influenza A/USSR/77 (H1N1)-like infection were given amantadine by small-particle aerosol with a Collison generator modified for this purpose. Inhalation periods for the volunteers were increased on consecutive weekends from 15 min to 1 h, 4 h, 9 h, and 2 consecutive days of 6 h each. The particle size was 1.2-micrometer mass median diameter, and the concentration of inhaled aerosol ranged from 47 to 75 microgram/liter. Estimates of retained doses in 9 h were 74 to 149 mg. About two-thirds of the dose was recovered in the urine. Pulmonary function studies did not vary significantly from base-line values and were within a normal range for five of seven volunteers. Two volunteers with a moderate reduction in mid-maximal flow before exposure had a total of three episodes of coughing and wheezing associated with moderate reductions in mid-maximal flow values. These episodes cleared spontaneously or improved promptly after isoproterenol therapy. The patients with influenza tolerated the treatment well and recovered promptly.

Adult

Effects of chronic amorphous silica exposure on sequential pulmonary function.

To assess clincial effects of precipitated amorphous silica (PAS), the authors reviewed serial spirograms, respiratory questionnaires, and chest radiographs of 165 workers exposed for a mean of 8.6 years. Monthly exposure was graded on a 1 to 4 scale and a "cumulative exposure index" (CEI) calculated for each worker from the sum of measured exposure. A "mean exposure index" (MEI) was calculated by dividing the CEI by total months exposed. Sputum production and dyspnea were inversely correlated with CEI, while cough and dyspnea correlated with mean pack-years of smoking but not PAS exposure. Linear regression analysis of yearly change of all pulmonary function variables (FVC, FEV1, FEV1/FVC, FEF25-75) showed no correlation with either the dose of PAS (CEI) or total years of exposure. Among 44 workers with a mean exposure time of 18 years (range 10-35 years), yearly decline of FVC and FEV1 were similar to the overall group. Of 143 workers with serial radiographs and exposure to only PAS, none had radiographic pneumoconiosis. Respiratory symptoms in PAS workers correlate with smoking but not with PAS exposure, while serial pulmonary function values and chest radiographs are not adversely affected by long-term exposure.

Adult

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Language

Clinical implications of a "nonspecific" transbronchial biopsy.

The diagnostic accuracy of transbronchial biopsy via fiberoptic bronchoscope was reviewed in 127 noncritically ill patients. Biopsy results were analyzed according to whether a "specific" pathologic diagnosis of neoplasm, granuloma or pneumonia, or a "nonspecific" diagnosis of inflammation, fibrosis or normal lung was made. The clinical significance of a "nonspecific" biospy specimen was evaluated by clinical follow-up of at least 12 months (mean 15 months) and by grouping patients according to the type of abnormality found on chest roentgenography. Clinical follow-up was available in 119 of these patients. The over-all "specific" diagnostic yield for biopsy with secretions was 49 per cent, with transbronchial biopsy being the sole means of specific diagnosis in 14 per cent of the patients with a peripheral mass lesion, in 18 per cent of the patients with localized infiltrative processes and in 52 per cent of the patients with diffuse infiltrative processes. In 64 (52 per cent) patients both biopsy specimens and secretions were diagnostically nonspecific. In 16 (77 per cent) patients with peripheral mass lesions but nonspecific biopsy findings and secretions, neoplasm was diagnosed by more invasive procedures. However, 22 (91 per cent) patients with localized and 12 (75 per cent) patients with diffuse infiltrative processes had benign clinical follow-up suggesting that open lung biopsy in such patients should be reserved for patients with obvious clinical or roentgenographic evidence of deterioration.

Biopsy

Evaluation of transbronchial biopsy of the lung.

Transbronchial biopsy of the lung through the fiberoptic bronchoscope is becoming a widely utilized clinical diagnostic technic. The authors reviewed the histologic features of 109 consecutive cases. The primary problem is the small size of the pulmonary tissue samples obtained, which is further compounded by an apparent mechanical inability to enter the biopsy forceps into dense nodules of neoplastic tissue. The diagnosis of carcinoma can best be made by examining the periphery of the cancer for microscopic extension into adjacent pulmonary parenchyma; however, multiple levels of the tissue block are needed to find such foci. A second problem reflects the use of transbronchial biopsy in the early clinical evolution of various infectious diseases in which pulmonary tissue may contain organisms without a fully developed inflammatory response, which would ordinarily provide the clue to the infective agent. The latter problem appears in cases of infections by Pneumocystis carinii. Close cooperation between the clinician and the pathologist is particularly important in all of these clinical situations.

Biopsy

An evaluation of several blood transfusion filters.

Several blood transfusion filters designed to remove microemboli were evaluated for filtration and flow characteristics, as well as for hemolysis and for release of particulate debris from the filter. The Dacron wool filter, a depth filter, was superior at removing microemboli measured with an electronic particle size analyzer. A 25 micrometer pore mesh filter was least effective at removing microemboli from stored blood, but demonstrated the most favorable flow rates and induced no apparent red blood cell destruction during filtration. In general, filter efficiency and filter induced hemolysis correlated inversely with flow rate. No filter tested released significant quantities of particulate debris when compared with unfiltered isotonic saline.

Blood Transfusion