Identification of vertical transmission of hepatitis B virus from mother to children by direct sequencing a segment of surface gene of hepatitis B virus

Intermittent catheterization (ICP) is a well-proven effective means of urologic management for spinal cord diseased (SCD) persons who meet the following criteria: adequate low pressure bladder capacity (350-400 cc minimum), adequate hand function, unobstructed urethra and compliant, understanding, continent, cooperative patients. Time-directed (Q4 H-Q6 H), ICP-obtained volumes on twenty-one patients revealed a majority of early, unnecessary as well as some late over-distended bladder catheterizations. The PCI 5000 or "Bladder Manager", a miniaturized ultrasonic bladder volume measuring device developed by Diagnostic Ultrasound of Seattle, was evaluated. It allowed the patients to perform volume-directed ICP which results in less frequent catheterizations and prevents bladder overdistension.     

Cloning, sequencing and expression of a cDNA encoding mammalian valyl-tRNA synthetase

The incus of the right ear from 4 growing mongrel dogs was surgically disarticulated and left in the middle ear space. The external auditory canal was then filled with teflon paste and sutured. After 6 weeks (D-6 group) and 13 weeks (D-13 group) the animals were sacrificed. The right experimental incus and the left control one were embedded in methyl methacrylate and sectioned in single 50-microns-thick sections according to the principal axis of the two processes. On the microradiographs of each section we evaluated the thickness of the body and of both processes and the percentage area of the primary channels of the secondary osteons and that of the appositional bone tissue. The thickness of the body and of the two processes was more pronounced in all the experimental incuses, in which 6% (in D-6) and 8% (in D-13) of the total area were occupied by new appositional woven bone. In the body of the D-13 group, 9% of the pre-existing bone was substituted by secondary osteons. The results indicate that the incus react to the variations of mechanical stimuli.     

cDNA cloning, sequence analysis, and induction by aryl hydrocarbons of a murine cytochrome P450 gene, Cyp1b1

C3H mouse embryo fibroblast cells, designated 10T1/2, can be transformed by physical and chemical agents including polycyclic aromatic hydrocarbons. In a previous report (Shen et al., Proc. Natl. Acad. Sci. USA 90, 11483-11487, 1993), we identified a cytochrome P450 gene induced by polycyclic aromatic hydrocarbons (PAHs) that is different from 1A1 or 1A2, and which we tentatively named P450CMEF. Here, we report the entire cDNA sequence of P450CMEF (5,128 bp) and the amino acid sequence deduced from it (543 residues). A comparison of the latter sequence with known cytochrome P450s indicates that P450CMEF is in a new subfamily of family 1 of the P450 superfamily. Accordingly, the Committee on Standardized Cytochrome P450 Nomenclature designated the gene Cyp1b1. Exposure to various aryl hydrocarbons (2.5 hr) induced Cyp1b1 mRNA in 10T1/2 cells to different degrees: 2,3,7,8-tetrachlorodibenzo-p-dioxin, 7,12-dimethylbenz[a]anthracene, benz[a]anthracene, benzo[a]pyrene, and beta-naphthoflavone were strong inducers; alpha-naphthoflavone and 3-methylcholanthrene, were moderate inducers; and benzo[e]pyrene was a weak inducer.     

Analysis of xylem formation in pine by cDNA sequencing

Secondary xylem (wood) formation is likely to involve some genes expressed rarely or not at all in herbaceous plants. Moreover, environmental and developmental stimuli influence secondary xylem differentiation, producing morphological and chemical changes in wood. To increase our understanding of xylem formation, and to provide material for comparative analysis of gymnosperm and angiosperm sequences, ESTs were obtained from immature xylem of loblolly pine (Pinus taeda L.). A total of 1,097 single-pass sequences were obtained from 5' ends of cDNAs made from gravistimulated tissue from bent trees. Cluster analysis detected 107 groups of similar sequences, ranging in size from 2 to 20 sequences. A total of 361 sequences fell into these groups, whereas 736 sequences were unique. About 55% of the pine EST sequences show similarity to previously described sequences in public databases. About 10% of the recognized genes encode factors involved in cell wall formation. Sequences similar to cell wall proteins, most known lignin biosynthetic enzymes, and several enzymes of carbohydrate metabolism were found. A number of putative regulatory proteins also are represented. Expression patterns of several of these genes were studied in various tissues and organs of pine. Sequencing novel genes expressed during xylem formation will provide a powerful means of identifying mechanisms controlling this important differentiation pathway.     

Cloning, sequencing and analysis of cDNA encoding bovine intercellular adhesion molecule-1 (ICAM-1)

Intercellular adhesion molecule-1 (ICAM-1) is an inducible glycoprotein that interacts with the leukocyte beta 2-integrins, LFA-1 and Mac-1. We have isolated and analyzed a cDNA clone coding for the putative bovine ICAM-1 gene and compared it with known comparative sequences from other species as well as bovine ICAM-3. The 3398-bp bovine ICAM-1 cDNA sequence codes for 535 amino acids and shows 57% homology with human ICAM-1 and 47% homology with bovine ICAM-3 at the amino acid levels. The predicted number and positions of cysteine residues in bovine ICAM-1 are all conserved among species including bovine ICAM-3. It has two arginine-glycine-aspartate (RGD) sites in the extracellular region and a serine residue in the cytoplasmic tail. Northern blot results show that the bovine ICAM-1 gene is expressed in stimulated leukocytes whereas bovine ICAM-3 is expressed predominantly in resting neutrophils.     

Functional studies with a full-length P-glycoprotein cDNA encoded by the hamster pgp1 gene

Hamster cells grown in culture may, like human and mouse cells, develop multidrug resistance (MDR) when exposed to certain cytotoxic chemotherapeutic agents. Several phenotypic features that are characteristic of MDR have been described; these include (1) resistance to many structurally and functionally unrelated drugs that have different cellular targets and modes of action; (2) reversal of MDR by certain agents, including verapamil and cyclosporin A; and (3) reduced intracellular drug accumulation relative to that of drug-sensitive cells. In this report we show that the introduction and overexpression of the hamster pgp1 cDNA confers to otherwise drug-sensitive cells an MDR phenotype with these features. Moreover, pgp1 transfectants showed varying degrees of resistance to anthracycline analogues, indicating that structural analogues of commonly used anticancer agents are capable of circumventing drug resistance conferred by pgp.     

Analysis of heterogeneous viral populations by direct DNA sequencing

The ability of direct PCR sequencing to detect and quantify sequence polymorphisms was investigated using samples containing mixed populations of HIV-1. A part of the genome encoding the polymorphic variable region 3 of the envelope was directly sequenced to yield a consensus sequence of the virus population. The results were compared with sequences obtained by analysis of multiple clones derived from the same clinical samples. The results of five patients suggested that the direct-sequencing method can be used as a rapid tool to analyze and quantify heterogeneous viral populations. Reconstitution experiments using cloned material demonstrated that it was possible to detect and quantify minor sequence variants present in as little as 10% of the total virus population. The use of the method for molecular diagnosis is discussed.     

Identification of a self-association region within the SCA1 gene product, ataxin-1

OBJECTIVES: To report the association between the protease inhibitor indinavir and the development of urolithiasis. METHODS: Case reports of three adult patients infected with the human immunodeficiency virus who developed surgical renal stones while being treated with indinavir are presented. RESULTS: Of the 3 patients requiring surgical intervention, stone analyses were available in 2. One stone revealed an inner core of an unidentifiable crystal surrounded by calcium oxalate, and another was found to have indinavir components as determined by thin-layer chromatography and gas chromatography-mass spectrometry. Metabolic evaluation of all 3 patients identified significant hypocitraturia as an isolated finding. CONCLUSIONS: The widely used protease inhibitor indinavir is associated with the development of urolithiasis and may act as a nidus for heterogeneous nucleation leading to the development of mixed urinary stones. Surgical intervention may be necessary in some cases. Underlying metabolic abnormalities may contribute to the increased incidence of stone formation. Urologists and other health care providers should be aware of this association, as combined medical and surgical intervention may be necessary.     

A novel oestrogen-regulated gene in human breast cancer cells identified by differential display

Shaker K+ channels inactivate through two distinct molecular mechanisms: N-type, which involves the N-terminal domain and C-type that appears to involve structural modifications at the external mouth of the channel. We have tested pore accessibility of the Shaker K+ channel during C-type inactivation using Ba2+ as a probe. We determined that external Ba2+ binds to C-type inactivated channels forming an extremely stable complex; i.e. there is Ba2+ trapping by C-type inactivated channels. The structural changes Shaker channels undergo during C-type inactivation create high energy barriers that hinder Ba2+ exit to either the extracellular solution or to the intracellular solution.     

Cloning, sequencing, and heterologous expression of rat methionine synthase cDNA

Methionine synthase catalyzes cobalamin-dependent methyl transfer reaction from 5-methyltetrahydrofolate to homocysteine, forming methionine. Rat methonine synthase cDNA was cloned and analyzed by RT-PCR, 3'- and 5'-RACE techniques. The cDNA consists of a 0.3-kb upstream untranslated region, a 3.8-kb coding region, and a 0.4-kb downstream untranslated region. The open reading frame encoded a polypeptide of 1,253 amino acid residues with a calculated molecular weight of 139,162. This molecular weight was in good agreement with the observed one (143,000) of the purified rat liver enzyme. The deduced amino acid sequence was 53, 92, and 64% identical with those of the Escherichia coli, human, and presumptive Caenorhabditis elegans enzymes, respectively. All the fingerprint sequences, forming parts of the cobalamin- and S-adenosylmethionine-binding sites, were completely conserved in the rat methionine synthase. A high-level expression of catalytically active enzyme in insect cells was done by infection with a baculovirus containing the rat methionine synthase cDNA.     

Detection of multiple hepatic micrometastases in pancreatic adenocarcinoma with a solitary liver metastasis by direct sequencing of the K-ras gene: a case report

Short-chain fructo-oligosaccharides (SC-FOS) are a mixture of oligosaccharides consisting of glucose linked to fructose units (Gfn; n = 相似文献   

VH and VL gene complexes encoding an anti-spectrin antibody are defined by nucleotide sequencing of cDNA from a hybridoma generated from Hu-PBL-SCID mouse

Previously we reported that human imunocompetent cells engrafted into scid mice mount a sustained and vigorous humoral immune response to murine erythrocytes. One of the dominant and consistently observed reactivity pattern of these antibodies in immunoblot analysis is with the alpha and beta isoforms of spectrin. In order to define the human xenoreactive response more completely, a hybridoma was generated (from a hu-PBL-scid mouse) whose antibody reacted with two high molecular weight species 225 to 250 kDa. We report here that this conserved antibody species reacts with both the murine and human erythrocyte proteins and cDNA nucleotide sequence analysis of the light and heavy chain genes encoding this antibody reveals that the light chain variable region gene has been previously observed in association with an autoreactive antibody. In addition to characterizing a conserved human B cell clonotype this is the first report of a human monoclonal antibody being generated from the hu-PBL-scid model using the standard hybridoma technology.     

Isolation of a candidate gene, CAB1, for cholesterol transport to mitochondria from the c-ERBB-2 amplicon by a modified cDNA selection method

An improved cDNA selection method was established to isolate expressed genes efficiently from an amplified chromosome region in human cancer. Biotinylated yeast artificial chromosome DNA containing c-ERBB-2 was hybridized in solution with PCR-amplifiable cDNAs of an esophageal cancer cell line bearing the c-ERBB-2 amplification. After capturing the hybrids on avidin-coated magnetic beads, the cDNAs were amplified by PCR. Four new genes (A39, C51, CAB1, and GRB-7) coamplified with c-ERBB-2 were isolated from the enriched cDNA library. CAB1, GRB-7, and c-ERBB-2 were overexpressed in gastric and esophageal cancer cells in correspondence with the amplification. The deduced amino acid sequence of the CAB1 gene had significant homology to the recently discovered steroidogenic acute regulatory protein, StAR, which plays an essential role in cholesterol transport to mitochondria. It was established that multiple overexpressed genes are frequently present in a single amplicon.     

Identification of a cDNA encoding 6-phosphogluconate dehydrogenase from a human heart cDNA library

A full-length cDNA clone for human 6-phosphogluconate dehydrogenase (PGD) was isolated from a human adult heart cDNA library. The clone encoded an open reading frame of 483 amino acids. When the amino acid sequences of human PGD and sheep PGD were aligned, 94.2% identity between these two proteins was found. Its calculated molecular weight is 53,149 daltons. The predicted isoelectric point is 6.85. When the secondary structure of human PGD was examined by the PROSIS software, 36% alpha-helix and 9% beta-sheet were found.     

Identification, cloning, and mutational analysis of the casein kinase 1 cDNA of the malaria parasite, Plasmodium falciparum. Stage-specific expression of the gene

The cDNA for casein kinase 1 (CK1) of Plasmodium falciparum was cloned, sequenced, and expressed in bacteria. The single major open reading frame of the 1.2-kilobase pair cDNA coded for a 324-amino acid polypeptide of approximately 37 kDa, the predicted sequence of which showed strong identity with known CK1 isoforms. The purified recombinant enzyme exhibited properties characteristic of CK1, such as inhibition by CK1-7, the ability to phosphorylate a highly specific peptide substrate, and a strong preference for ATP over GTP. A casein kinase activity, partially purified from soluble extracts of P. falciparum by affinity chromatography through CK1-7 columns displayed identical properties. The activity showed a stage-specific expression in the parasite, in the order trophozoite > ring > schizont. Northern analysis indicated the existence of two major CK1 mRNAs, 2.4 and 3.2 kilobase pairs long, the levels of which were in the order ring > schizont > trophozoite. Mutagenesis of recombinant CK1 defined important amino acid residues and their potential role in the conformation of the enzyme. The malarial CK1 appeared to be the one of the smallest and perhaps the most primitive CK1 enzymes known, containing little sequence information beyond the minimal catalytic domain.