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Biomedical subjects

C Boyd

Publications and source records attributed to C Boyd.

At least 19 recordsLinked to original sources

Cloning and characterization of a phospholipase gene from Erwinia chrysanthemi EC16.

A single gene (plcA) was cloned from a cosmid library of Erwinia chrysanthemi EC16 DNA that encoded an extracellular phospholipase. The gene was subcloned and DNA sequence data showed the presence of a single open reading frame encoding a protein with a predicted size of 39 kDa. The coding region was G+C-rich and the protein had a predicted basic isoelectric point. The protein showed no significant homology with others in the PIR library, including other phospholipases. When overexpressed in Escherichia coli cells, the plcA gene directed production of a c. 39 kDa protein that was largely localized in the periplasm, but its N-terminal amino acid sequence was that of the native protein predicted from DNA sequence data. Unlike the wild-type bacterium, an E. chrysanthemi EC16 marker exchange mutant of the plcA gene did not secrete extracellular phospholipase activity in the medium. However, no detectable change was observed in terms of the virulence of the mutant strain on potato tubers or chrysanthemum stems.

Amino Acid Sequence

Homograft valve durability: host or donor influence?

Antibiotic sterilized valves have been shown to function longer than those chemically sterilized; however, the reason remains obscure. Current hypotheses cite either retention of donor fibroblasts capable of repairing the grafted valve, or host fibroblast ingrowth into and onto the leaflet ground substance. A cryopreserved aortic homograft from a male donor was explanted from a female recipient after 10 months, and subjected to immunocytochemistry, tissue culture, and karyotyping. The leaflet bases exhibited normal morphology with an intact endothelium. The distal one-third of the leaflets was devoid of fibroblasts from the leaflet bases showed them to be of host origin. This homograft seems to have been implanted with an intact ground substance which allowed for host cell repopulation of the inner one-third of the leaflets. Perhaps donor cell viability in itself is not as important to durability as is preservation of the leaflet ground substance, but rather the presence of viable cells may be an index of the structural integrity of the collagen and elastic matrix.

Adult

Luteal phase deficiency after completely normal follicular and periovulatory phases.

Luteal phase defect (LPD) accounts for a significant proportion of reproductive disorders, however its etiology is still debated. A prospective study was performed on 37 ovulatory women to determine whether LPD can occur in cycles characterized by completely normal folliculogenesis. Criteria for normal folliculogenesis included: a gradual rise of serum estradiol, a luteinizing hormone (LH) surge, the presence of a dominant follicle that disappeared, an increase of serum progesterone, and normal serum levels of prolactin, testosterone, dehydroepiandrosterone sulfate, follicle-stimulating hormone, and LH. Thirty of 37 women fulfilled the above mentioned strict criteria and underwent endometrial biopsy in the late luteal phase. Seven of 30 (23%) demonstrated a delay in endometrial development and all had normal hormonal and ultrasonographic parameters of folliculogenesis and ovulation. Women with delayed endometrial development demonstrated slightly longer follicular phases (17.0 +/- 1.1 versus 14.5 +/- 0.3 days). Perfectly normal follicular and periovulatory events may be followed by deficient luteal phases.

Adult

Structure, polymorphism, and novel repeated DNA elements revealed by a complete sequence of the human alpha-fetoprotein gene.

The human alpha-fetoprotein gene spans 19,489 base pairs from the putative "Cap" site to the polyadenylation site. It is composed of 15 exons separated by 14 introns, which are symmetrically placed within the three domains of alpha-fetoprotein. In the 5' region, a putative TATAAA box is at position -21, and a variant sequence, CCAAC, of the common CAT box is at -65. Enhancer core sequences GTGGTTTAAAG are found in introns 3 and 4, and several copies of glucocorticoid response sequences AGATACAGTA are found on the template strand of the gene. There are six polymorphic sites within 4690 base pairs of contiguous DNA derived from two allelic alpha-fetoprotein genes. This amounts to a measured polymorphic frequency of 0.13%, or 6.4 X 10(-4)/site, which is about 5-10 times lower than values estimated from studies on polymorphic restriction sites in other regions of the human genome. There are four types of repetitive sequence elements in the introns and flanking regions of the human alpha-fetoprotein gene. At least one of these is apparently a novel structure (designated Xba) and is found as a pair of direct repeats, with one copy in intron 7 and the other in intron 8. It is conceivable that within the last 2 million years the copy in intron 8 gave rise to the repeat in intron 7. Their present location on both sides of exon 8 gives these sequences a potential for disrupting the functional integrity of the gene in the event of an unequal crossover between them. There are three Alu elements, one of which is in intron 4; the others are located in the 3' flanking region. A solitary Kpn repeat is found in intron 3. The Xba and Kpn repeats were only detected by complete sequencing of the introns. Neither X, Xba, nor Kpn elements are present in the related human albumin gene, whereas Alu's are present in different positions. From phylogenetic evidence, it appears that Alu elements were inserted into the alpha-fetoprotein gene at some time postdating the mammalian radiation 85 million years ago.

Base Sequence

Reye's syndrome.

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Female

Ejection fraction by combined inverse Fourier analysis and second-derivative technique: correlation with isocontour method.

In order to measure ejection fractions (EFs) from nuclear ventriculograms, we devised a semi-automated edge-detection technique based on a combination of inverse Fourier analysis and second-derivative techniques. Initial clinical studies showed that, for the left ventricle, our method gives EF values statistically identical with those obtained using a conventional isocontour technique. For the right ventricle, however, the values obtained using the two methods were somewhat more at variance. Despite requiring a longer processing time, the results obtained with our method are reproducible because less operator intervention is necessary.

Fourier Analysis

Mutations linked to the pro alpha 2(I) collagen gene are responsible for several cases of osteogenesis imperfecta type I.

We have analysed six South African families with osteogenesis imperfecta type I using three DNA polymorphisms associated with the pro alpha 2(I) collagen gene. In four of these families linkage of the pro alpha 2(I) gene and the osteogenesis imperfecta phenotype was suggested, whereas in the remaining two families there was a lack of linkage. No distinct correlation could be made between the phenotypic features of the families studied and linkage or lack of linkage to the pro alpha 2(I) gene. Two different haplotypes were found to be associated with the mutant pro alpha 2(I) alleles. These findings suggest that molecular heterogeneity exists within osteogenesis imperfecta type I and that in a significant proportion of cases the defect is linked to the pro alpha 2(I) gene.

DNA

Structure of the 3' portion of the bovine elastin gene.

A bovine genomic library constructed by partial Sau3A digestion and contained in lambda Charon 30 was screened by in situ hybridization with a 1.3-kilobase (kb) sheep elastin cDNA clone [Yoon, K., May, M., Goldstein, N., Indik, Z., Oliver, L., Boyd, C., & Rosenbloom, J. (1984) Biochem. Biophys. Res. Commun. 118, 261-269]. Three clones encompassing 10 kb of the bovine elastin gene were identified and characterized by restriction mapping and DNA sequencing of the 6.2 kb of the most 3' region of the gene. These analyses have permitted localization of eight exons in the 6.2 kb in which the translated exons vary in size from 27 to 69 base pairs, and there is an approximately 1-kb untranslated region at the 3' end. In addition to identification of sequences homologous to those found in porcine tropoelastin, the analyses defined a 58 amino acid sequence that forms the carboxy-terminal region of tropoelastin, and this sequence, which contains two cysteine residues, was previously not observed in the protein sequence data. The analyses also suggest that functionally distinct cross-link and hydrophobic domains of the protein are encoded in separate exons.

Amino Acid Sequence

Analysis of the 3' region of the sheep elastin gene.

The nucleotide sequences of a 1279-bp sheep elastin cDNA clone, pcSEL1 [Yoon et al. (1984) Biochem. Biophys. Res. Commun. 118, 261-269], and a 1230-bp sheep elastin genomic subclone, pSS1 [Davidson et al. (1984) Biochem. J. 220, 643-652], corresponding to a portion of the cDNA clone, were determined. These analyses permitted determination of the 100 amino acids at the carboxy terminus of sheep tropoelastin. A portion of this sequence showed strong homology to known sequences of pig tropoelastin, but most of the sequence had not been previously determined through protein sequencing. Novel aspects of the tropoelastin molecule which have been revealed by the present analyses are (i) the presence of an unusual sequence, KPPKP, which may contribute to crosslink formation; and (ii) the finding of cysteine within a sequence, CLGKSCGRKRK, at the putative carboxy terminus of tropoelastin. Because of the presence of these sequences, it is speculated that the carboxy-terminal region may be of importance in crosslinking tropoelastin molecules to themselves or to other matrix macromolecules. The nucleotide analyses revealed that sheep elastin mRNA contains a 974-bp untranslated sequence at the 3' end, which appears to be strongly conserved among species.

Animals

Toward an understanding of mother-daughter identification using concept analysis.

Mother-daughter identification was analyzed using a concept analysis strategy proposed by Walker and Avant. This strategy was selected because of its utility in clarifying a concept requiring refinement. It was toward this goal that the concept analysis of mother-daughter identification was initiated. Rogers's conceptual model was used to add a new dimension to past conceptualizations of mother-daughter identification, one that allows the concept to be added to a nursing perspective.

Adult

Chromosomal localization of the human elastin gene.

mRNA isolated from fetal human aorta was used to synthesize cDNA that was cloned into the PstI site of pBR322. The recombinant clones were screened with an authentic sheep elastin cDNA, and one human clone that hybridized strongly was isolated and characterized. The 421-base pair (bp) insert of this human clone was sequenced by the dideoxy method, and the DNA sequence showed strong homology to the nontranslated portion of the sheep elastin cDNA. This result unequivocally identified the human clone, designated pcHEL1, as an elastin clone. Plasmid pcHEL1 labeled with [3H] nucleotides was used in in situ hybridization experiments utilizing normal metaphase chromosomes and also with cells carrying a balanced translocation between chromosomes 1 and 2: 46,XY,t(1;2)(p36;q31). The results strongly suggest that the elastin gene is localized to the q31----qter region of chromosome 2.

Animals

Disappearance of Ph1-positive cells in four patients with chronic granulocytic leukemia after chemotherapy, irradiation and marrow transplantation from an identical twin.

Four patients (21, 41, 13 and 38 years of age) with a history of chronic granulocytic leukemia for 12, 10, 11, and 106 months, respectively, were treated with dimethyl busulfan, cyclophosphamide, 920 rads of total-body irradiation and intravenous marrow infusion from normal, genetically identical twins. Serial chromosome analyses were performed on marrow aspirates cultured without mitotic stimulants. No Ph1-positive cells were detected in the marrows from the normal twins, whereas just before therapy, all 100 metaphases examined from each patient were Ph1-positive. Chromosome analyses were performed three to five times per patient after transplantation, and not a single Ph1-positive cell was detected. The patients remain hematologically normal 22, 23, 26 and 31 months after transplantation. The results show that the Ph1-positive clone can be eradicated by vigorous therapy and that the marrow in chronic granulocytic leukemia can be repopulated by stem cells from normal twins.

Adult