RESUMO
In benign and malignant ovarian tumor patients, human placental alkaline phosphatase (HPLAP) was determined in serum and extracts from surgical tumor biopsies using a highly specific enzyme-antigen immunoassay based on a mouse monoclonal antibody (E6) to HPLAP. Serum HPLAP levels greater than or equal to 0.1 unit/liter were found in 58% of ovarian cancer patients. Serum carcinoembryonic antigen levels were positive (greater than 5.4 ng/ml) in 17% of these patients. HPLAP was detected in extracts from 13 of the 14 tumors investigated (range, 2.4 to 557 milliunits/g). Only the mixed heterologous Müllerian sarcoma was negative. The highest HPLAP content of normal ovarian tissue was 1.1 milliunits/g. The amount of heat-stable and L-p-bromotetramisole-insensitive alkaline phosphatase was in all cases much higher than the fraction recognized by E6. The neoplastic origin of HPLAP was confirmed immunohistochemically on paraffin sections by an indirect avidin-biotin-peroxidase staining procedure using E6. The staining pattern was compared to the histochemical distribution of total alkaline phosphatase on adjacent sections. A consistency was found between the amount of HPLAP in tissue extracts and its immunohistochemical distribution. In all the tumors, staining for HPLAP was observed mainly on the plasma membranes of carcinoma cells. In 9 of the 10 carcinomas, the histological distribution of HPLAP and also of total alkaline phosphatase was heterogeneous. HPLAP staining, present in one of five normal ovaries, was restricted to germinal inclusion cysts. The present results support the hypothesis that serous ovarian tumors originate from these cysts.
Assuntos
Fosfatase Alcalina/análise , Isoenzimas/análise , Neoplasias Ovarianas/enzimologia , Antígeno Carcinoembrionário/análise , Eletroforese , Feminino , Histocitoquímica , Humanos , Neoplasias Ovarianas/patologia , Placenta/enzimologia , GravidezRESUMO
Children acquire neuropsychologic dysfunctions after chemotherapy for hematologic malignancy. In this study, putative changes in levels of CSF-tau (a marker of neural dysintegrity) in leukemic children prior to and during chemotherapy were studied. Cerebrospinal fluid (CSF) samples were obtained before and during treatment from patients with B cell non-Hodgkin's lymphoma (NHL, n = 10), non-B cell acute lymphoblastic leukemia/NHL (non-B-ALL, n = 48), acute myeloid leukemia (AML, n = 9), other malignant diseases (n = 9), and six control children. A sandwich-type ELISA (INNOTEST hTAU-Ag) was used for measuring CSF-tau. Sixteen out of 50 patients with hematological malignancies, including the patients with proven leukemic CNS invasion, already showed high CSF-tau levels at baseline (>300 pg/ml). The pre-induction treatment for non-B-ALL, consisting of only corticosteroids and methotrexate (MTX), resulted in a significant increase of tau at day 8 (on average to 535 pg/ml). Larger increases as compared to baseline levels of CSF-tau were observed in patients treated for B-NHL with systemic vincristine, corticosteroids and cyclophosphamide, and intrathecal MTX (mean 776 pg/ml at day 8). In two AML patients with CNS invasion, CSF-tau increased during chemotherapy up to 1,500 and 948 pg/ml, respectively. In one non-B-ALL patient with MTX-induced clinical neurotoxicity, CSF-tau was above the detection limit of 2,000 pg/ml. Almost one-third of the patients with hematological malignancies had elevated CSF-tau levels at diagnosis. Transient high levels of CSF-tau, reaching levels observed in other neurodegenerative disorders, were observed during induction chemotherapy for non-B-ALL, B-NHL and CNS+ AML. The clinical implications of both observations will be the subject of further study.
Assuntos
Antineoplásicos/efeitos adversos , Biomarcadores/líquido cefalorraquidiano , Neoplasias Hematológicas/líquido cefalorraquidiano , Proteínas tau/líquido cefalorraquidiano , Antineoplásicos/uso terapêutico , Criança , Neoplasias Hematológicas/tratamento farmacológico , Humanos , Testes NeuropsicológicosRESUMO
Roof maintenance practices often involve the application of biocide products to fight against moss, lichens and algae. The main component of these products is benzalkonium chloride, a mixture of alkyl benzyl dimethyl ammonium chlorides with mainly C12 and C14 alkyl chain lengths, which is toxic for the aquatic environment. This paper describes, on the basis of an in-situ pilot scale study, the evolution of roof runoff contamination over a one year period following the biocide treatment of roof frames. Results show a major contamination of roof runoff immediately after treatment (from 5 to 30 mg/L), followed by an exponential decrease. 175-375 mm of cumulated rainfall is needed before the runoff concentrations become less than EC50 values for fish (280 µg/l). The residual concentration in the runoff water remains above 4 µg/L even after 640 mm of rainfall. The level of benzalkonium ions leaching depends on the roofing material, with lower concentrations and total mass leached from ceramic tiles than from concrete tiles, and on the state of the tile (new or worn out). Mass balance calculations indicate that a large part of the mass of benzalkonium compounds applied to the tiles is lost, probably due to biodegradation processes.
Assuntos
Compostos de Benzalcônio/análise , Materiais de Construção , Desinfetantes/análise , Chuva , Poluentes Químicos da Água/análise , Monitoramento Ambiental , França , Projetos Piloto , Movimentos da ÁguaRESUMO
We report here the synthesis, in nonlymphoid cells, of two functionally active recombinant F(ab')2 fragments directed against the tumor marker, human placental alkaline phosphatase (hPLAP). The truncated heavy chain (HC) sequences, E6Hf2 and E6Hy3f2, of the murine F(ab')2 fragment, E6F2, and of the murine::human chimeric F(ab')2 fragment, E6(Hy3,kappa)F2, respectively, were engineered by introducing an in-phase stop codon within the second constant domain of the corresponding parental HC sequence. The antibody-encoding genes were placed under control of the simian virus 40 late promoter and each HC sequence, together with the light chain (LC) sequence, was transiently expressed in COS-1 cells. The truncated HCs were correctly synthesized, processed and assembled with the murine LC and subsequently secreted into the culture medium as functionally active entities with stable hinge region interactions. These results indicate that, under the conditions used, the hinge region was sufficient for the formation of divalent molecules. However, Western blotting revealed the presence of hPLAP-binding half-molecules of E6F2, which was not the case for E6(Hy3,kappa)F2. Since E6F2 and E6(Hy3,kappa)F2 mainly differ by the length of their hinge region (22 and 62 aa residues, respectively) and the number of inter-HC disulfide bridges (four and eleven, respectively), it may be concluded that F(ab')2 fragments with an extended hinge region and several inter-HC disulfide bridges are formed more efficiently.
Assuntos
Quimera , Fragmentos Fab das Imunoglobulinas/genética , Fosfatase Alcalina/imunologia , Fosfatase Alcalina/metabolismo , Animais , Sequência de Bases , Western Blotting , Linhagem Celular , Eletroforese em Gel de Poliacrilamida , Feminino , Expressão Gênica , Humanos , Fragmentos Fab das Imunoglobulinas/metabolismo , Camundongos , Dados de Sequência Molecular , Placenta/enzimologia , Plasmídeos , Gravidez , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , TransfecçãoRESUMO
IgG1-secreting variants have been isolated from three different IgM-secreting hybridomas, in two instances following in vitro immunization. The method used was based on sequential sublining in combination with selection by an IgG1-specific two-site ELISA system employing two different IgG1-specific polyclonal antisera. Idiotypic identity between the IgG1 variants and their respective IgM parent was demonstrated using syngeneic anti-idiotypic antisera. The antigen binding specificity in the IgG1 variants was also conserved. Isolation of naturally occurring IgG1 switch variants from IgM-secreting hybridomas that are produced after in vivo immunization offers a solution to the major disadvantages associated with the generation of IgM hybridomas.
Assuntos
Hibridomas/imunologia , Imunoglobulina G/isolamento & purificação , Imunoglobulina M/imunologia , Doença de Alzheimer/metabolismo , Animais , Anticorpos Anti-Idiotípicos/imunologia , Encéfalo/metabolismo , Eletroforese em Gel de Poliacrilamida , Ensaio de Imunoadsorção Enzimática , Humanos , Técnicas Imunoenzimáticas , Imunoglobulina G/imunologia , Região de Troca de Imunoglobulinas , Camundongos , Camundongos Endogâmicos BALB C , Neurofibrilas/metabolismoRESUMO
Hydrogel films, prepared by cross-linking of gelatin with dextran dialdehydes (weight ratio 2:1), and containing either fluorescein isothiocyanate dextran (Mw 70000) or polypeptides were evaluated in terms of their release characteristics and mechanical properties upon increasing storage time at 4 degrees C. Important changes in release kinetics and mechanical properties of the cross-linked gelatin films were observed, especially during the first week after the hydrogel production. Rheological and NMR measurements showed that the mechanical properties of the gelatin hydrogel films were improved with increasing storage time. It appeared that the process of chemical cross-linking and physical structuring of the gelatin hydrogel matrix did not occur instantaneously and substantially influenced the polypeptide release patterns. Cross-linked gelatin hydrogels were found to be appropriate release systems for medium-term sustained delivery of biologically active epidermal growth factor (EGF), but release characteristics were strongly dependent on the nature of the protein which was incorporated.
Assuntos
Reagentes de Ligações Cruzadas/química , Dextranos/química , Gelatina/química , Polietilenoglicóis/química , Proteínas/química , Animais , Células Cultivadas , Fenômenos Químicos , Físico-Química , Preparações de Ação Retardada , Elasticidade , Fator de Crescimento Epidérmico/administração & dosagem , Fator de Crescimento Epidérmico/química , Fator de Crescimento Epidérmico/farmacocinética , Fluoresceína-5-Isotiocianato/análogos & derivados , Fluoresceína-5-Isotiocianato/química , Hidrogel de Polietilenoglicol-Dimetacrilato , Radioisótopos do Iodo , Cinética , Camundongos , Camundongos Endogâmicos BALB C , Proteínas/administração & dosagem , Proteínas/farmacocinética , Soroalbumina Bovina/administração & dosagem , Soroalbumina Bovina/química , Soroalbumina Bovina/farmacocinética , ViscosidadeRESUMO
The biosafety of a new hydrogel wound dressing material consisting of dextran dialdehyde cross-linked gelatin was evaluated (i) in vitro in cultures of dermal fibroblasts, epidermal keratinocytes, and endothelial cells, three cell types which play a major role in the process of cutaneous wound healing, and (ii) in vivo by subcutaneous implantation studies in mice. The cytotoxicities of this hydrogel, two semi-occlusive polyurethane dressings (Tegaderm and OpSite), and a hydrocolloid dressing (DuoDERM) were compared by measuring cell survival with the tetrazolium salt reduction (MTT) assay after incubations of the wound dressing samples for up to 6 d, in the presence of--but not in direct contact with--the cells. In vitro, the degree of cytotoxicity of the new hydrogel was greater in keratinocyte cultures than in fibroblast and endothelial cell cultures, and increased upon longer incubation time. In keratinocyte cultures, the semi-occlusive polyurethane dressings, the hydrocolloid, and the hydrogel dressings induced low, high and acceptable degrees of cytotoxicity, respectively. The toxicity of the isolated hydrogel components was assessed in Balb MK keratinocyte cultures. In these cells, epidermal growth-factor-stimulated thymidine incorporation into DNA was higher in the presence of gelatin. By contrast, concentrations of dextran dialdehyde as low as 0.002% were found to significantly decrease thymidine incorporation (P < 0.01). Subcutaneous implantation studies in mice showed that in vivo the hydrogel was biocompatible since the foreign body reaction seen around the implanted hydrogel samples was moderate and became minimal upon increasing implantation time. These results indicate that dextran dialdehyde cross-linked gelatin hydrogels have an appropriate biocompatibility.
Assuntos
Materiais Biocompatíveis/toxicidade , Reagentes de Ligações Cruzadas/toxicidade , Dextranos/toxicidade , Gelatina/toxicidade , Hidrogel de Polietilenoglicol-Dimetacrilato/toxicidade , Implantes Experimentais , Células 3T3/efeitos dos fármacos , Animais , Materiais Biocompatíveis/química , Células Cultivadas , Reagentes de Ligações Cruzadas/química , Dextranos/química , Endotélio/citologia , Endotélio/efeitos dos fármacos , Gelatina/química , Humanos , Hidrogel de Polietilenoglicol-Dimetacrilato/química , Queratinócitos/citologia , Queratinócitos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Cell cultures of primary mouse granulosa cells were transfected with a v-myc-containing plasmid, and the resulting stable cell lines were tested for their steroidogenic properties and physiologic status. Granulosa cells were obtained from 22-day-old NMRI mice injected with 8 IU pregnant mare serum gonadotropin i.p. 2 days earlier. In Passage 1 the cells were transfected with pSVv-myc using calcium phosphate precipitation or lipofectin. The 3 beta- and 17 beta-hydroxy steroid dehydrogenase activity was visualized in control cultures. The three cell lines obtained have been in culture for over 1 yr and have been subcultured for more than 90 passages. The cell line GRM01, with a doubling time of 37 +/- 3 h and a diploid modal chromosome number, produced progesterone, estradiol, as well as inhibinlike and activinlike material under basal conditions. A combination of follicle-stimulating hormone and luteinizing hormone was able to increase the secretion of progesterone. GRM01L, a fast growing clone of the GRM01 line with a doubling time of 10 +/- 1 h, retained only the capacity to produce activinlike material and transforming growth factor-beta, and it was the only one with a tumorigenic capacity. Epidermal growth factor, insulin, and interleukin-6 were able to induce the [3H]thymidine incorporation into DNA in these two cell lines. GRM02, with a doubling time of 36 +/- 2 h and a hypertriploid modal chromosome number, produced progesterone and activinlike and inhibinlike material. Follicle-stimulating hormone and luteinizing hormone were able to enhance the secretion of progesterone. For this cell line, only insulin was shown to induce [3H]thymidine incorporation into DNA.
Assuntos
Linhagem Celular Transformada/fisiologia , Células da Granulosa/fisiologia , 17-Hidroxiesteroide Desidrogenases/metabolismo , 3-Hidroxiesteroide Desidrogenases/metabolismo , Ativinas , Animais , Divisão Celular/efeitos dos fármacos , Linhagem Celular Transformada/química , Linhagem Celular Transformada/citologia , Meios de Cultivo Condicionados , Feminino , Células da Granulosa/química , Células da Granulosa/citologia , Substâncias de Crescimento/farmacologia , Inibinas/metabolismo , Cariotipagem , Camundongos , Prostaglandinas/metabolismo , Proteínas Proto-Oncogênicas c-myc/análise , Esteroides/análise , Fator de Crescimento Transformador beta/metabolismoRESUMO
Neo natal androgenisation of the female rat provoke at 3 months : 1) an accentuation of the insulin secretion at the glucose loading with the diet rich in saturated fat an a reduction with the diet rich in sucrose or rapesced oil; 2) Hypercholesterolemia with all the diets and 3) hypertriglyceridemia with diet rich in saturated fat or sucrose. Neo natal oestrogenisation of the male rata provoke at 3 months : 1) a reduction of the insulin secretion at the glucose loading with all the diets 2) hypercholesterolemia with the diet rich in saturated fats 3) hypertriglyceridemia with diets rich in M.C.T. or sucrose.
Assuntos
Estradiol/farmacologia , Testosterona/farmacologia , Animais , Animais Recém-Nascidos , Colesterol/sangue , Colesterol na Dieta/administração & dosagem , Feminino , Hipercolesterolemia/induzido quimicamente , Hiperlipidemias/induzido quimicamente , Insulina/metabolismo , Secreção de Insulina , Masculino , Ratos , Triglicerídeos/administração & dosagem , Triglicerídeos/sangueAssuntos
Anticorpos Monoclonais/uso terapêutico , Antígeno B7-1/imunologia , Ciclosporina/uso terapêutico , Rejeição de Enxerto/prevenção & controle , Transplante de Pele/imunologia , Linfócitos T/imunologia , Animais , Rejeição de Enxerto/imunologia , Sobrevivência de Enxerto/efeitos dos fármacos , Sobrevivência de Enxerto/imunologia , Teste de Histocompatibilidade , Terapia de Imunossupressão/métodos , Macaca mulatta , Linfócitos T/efeitos dos fármacosRESUMO
The relative contribution of the transmembrane segments in the alpha-subunit of Shaker-type potassium channels was investigated in relation to potassium channel function. Starting from a wild-type Kv1.1 channel, four different deletion mutants were made, missing respectively transmembrane segments S1 and S2, S2 and S3, S1 to S3, and S1 to S4. To ensure the assembly of the different subunits, the hydrophylic N-terminal domain was always conserved. The lack of transmembrane segments S1 to S4 converts a depolarization-activated WT Kv1.1 channel with outward rectification into a hyperpolarization-activated channel with inward rectification. In contrast, mutant channels missing transmembrane segments S1 and S2, S2 and S3, or S1 to S3 did not reveal functional expression.
Assuntos
Oócitos/fisiologia , Peptídeos/fisiologia , Canais de Potássio/fisiologia , Deleção de Sequência , Animais , Membrana Celular/fisiologia , Membrana Celular/ultraestrutura , Clonagem Molecular , DNA Complementar , Drosophila , Expressão Gênica , Globinas/biossíntese , Potenciais da Membrana , Modelos Estruturais , Mutagênese Sítio-Dirigida , Biossíntese Peptídica , Canais de Potássio/biossíntese , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/metabolismo , Mapeamento por Restrição , Superfamília Shaker de Canais de Potássio , XenopusRESUMO
The nucleotide sequence of the simian virus 40 (SV40) genome region between the cleavage sites for restriction endonucleases EcoRI (map position 0) and HindII (map position 0.05) has been determined mainly by the partial chemical DNA degradation procedure of Maxam and Gilbert. This fragment represents 5.3% of the genome of SV40 and is located in the late region, internally in the VP1 gene. The message strand shows only one open reading frame for translation into protein, which connects to the one for the preceding fragment. On this basis part of the amino acid sequence of the VP1 protein is presented.
Assuntos
Enzimas de Restrição do DNA/metabolismo , DNA Viral/análise , Vírus 40 dos Símios/análise , Sequência de Bases , Códon , Escherichia coli/enzimologia , Genes Virais , Conformação de Ácido Nucleico , RNA Mensageiro/análise , Proteínas Virais/biossínteseRESUMO
The restriction fragment Hind-G represents 7.0% of the simian virus 40 (SV40) genome. The information present in fragment Hind-G is expressed as part of the major, late 16-S messenger RNA. The complete nucleotide sequence of the fragment Hind-G has now been determined by application of the procedure of Maxam and Gilbert [Proc. Natl Acad. Sci. U.S.A. (1977) 74, 560-564]. It contains 369 nucleotide base pairs. On the basis of the termination code words in the strand with the same polarity as the late mRNA, two illegitimate reading frames can be defined. Therefore the third, open frame must code for the carboxyl terminal part of the VP1 protein. It terminates within fragment Hind-G with a TGA signal. This stop codon is followed by a non-translated region of the mRNA of about 83 nucleotides. The latter contains the sequence A-A-U-A-A-A, common to all other eukaryotic mRNA molecules so far studied. The Hind-G fragment also contains sequences which presumably play a role in the synthesis, processing and/or expression of early mRNA; these aspects are discussed in the following paper.
Assuntos
Códon , DNA Viral/análise , RNA Mensageiro , Vírus 40 dos Símios , Proteínas Virais/biossíntese , Aminoácidos , Sequência de Bases , Enzimas de Restrição do DNA/metabolismo , Escherichia coli/enzimologiaRESUMO
The HindII + III restriction fragment J (Hind-J) represents 4.58% of the simian virus 40 genome. The information present in Hind-J is expressed as part of the major, late 16-S messenger RNA, which codes for the structural protein VP1. The nucleotide sequence of the 240-base-pairs-long Hind fragment J has been determined by analysis of each oligonucleotide from both strands resulting from T1 or pancreatic RNase digestion of RNA transcribed from the DNA and from RNase digestion of ribo-substituted DNA. Large oligonucleotide blocks which could be constructed mainly on the basis of complementarity were subsequently ordered by partial chemical degradation of terminally labeled DNA. This direct DNA sequencing approach also completely confirmed the results obtained by both aforementioned RNase degradation methods. In the strand with the same polarity as the late mRNA, triplets corresponding to termination codons are present in two of the three reading frames. The one open reading frame connects in phase with the open reading frame of the neighboring HindII/ III fragments K, F and G, which have been published previously and which together with Hind-J span the total VP1 gene. Some features of the primary nucleotide sequence of this VP1 gene and the derived VP1 amino acid sequence are discussed.
Assuntos
DNA Viral , Genes Virais , Vírus 40 dos Símios/análise , Proteínas Virais , Sequência de Aminoácidos , Aminoácidos/análise , Sequência de Bases , Enzimas de Restrição do DNA , Haemophilus influenzae/enzimologia , Oligodesoxirribonucleotídeos/análise , RNA MensageiroRESUMO
The nucleotide sequence of the second part of the simian virus 40 DNA HindII + III restriction fragment A is presented. The sequence extends from map position 0.533 to 0.424 and together with the first part of Hind-A [Volckaert et al. Proc. Natl Acad. Sci. U.S.A. 75, 2160--2164 (1978)] completes the total hind-A sequence, comprising 1169 base pairs. The second half of Hind-A includes the region corresponding to the second splicing boundary common to small tumor antigen (small-t) and large tumor antigen (large-T) mRNA and it contains coding information for an internal portion of large-T antigen. Two similar secondary structures of reasonable thermodynamic stability can be proposed for the nucleotide sequence of the pre-mRNA corresponding to the region reported here. Their possible relevance to the splicing of the SV40 early mRNAs is discussed. The deduced amino acid sequence is 188 residues long and contains a Lys-Lys-Lys-Arg-Lys-stretch which may be involved in the DNA binding capacity of large-T. A presumptive phosphorylation site is also present.
Assuntos
Antígenos Virais/genética , DNA Viral , Desoxirribonucleases de Sítio Específico do Tipo II , Genes Virais , Vírus 40 dos Símios/genética , Sequência de Aminoácidos , Antígenos Virais de Tumores , Sequência de Bases , Enzimas de Restrição do DNA , Desoxirribonuclease HindIII , Conformação de Ácido NucleicoRESUMO
The HindII + III restriction fragment I (Hind-I) from simian virus 40 DNA represents 4.96% of the genome and maps in the early transcription region. Hind-I is an internal segment of the A gene and its information is expressed as part of the early 19-S mRNA, which codes for T antigen. We report here the nucleotide sequence of the 259-base-pair Hind-I fragment. The sequence was determined and confirmed by RNA and DNA sequencing methods: by analysis of oligonucleotides resulting from T1 and pancreatic RNase digestion of labeled RNA transcribed from SV40 DNA with Escherichia coli DNA-dependent RNA polymerase, by partial degradation of RNA transcripts with snake venom phosphodiesterase, and by base-specific chemical degradation of 5'-end-labeled subfragments of Hind-I according to the procedure of Maxam and Gilbert. Multiple triplets corresponding to termination codons occur in two of the three reading frames of the DNA strand that has the same polarity as early mRNA. The open reading frame connects in phase with the one of the Hind fragments flanking Hind-I, and the amino acid sequence specified by Hind-I lies in the middle part of the large-T antigen. Some features of the primary nucleotide sequence and of early transcription are discussed.
Assuntos
Antígenos Virais/genética , DNA Viral , Desoxirribonucleases de Sítio Específico do Tipo II , Genes Virais , Vírus 40 dos Símios/genética , Antígenos Virais de Tumores , Sequência de Bases , Enzimas de Restrição do DNA , Desoxirribonuclease HindIII , Oligorribonucleotídeos , RNA Viral , Transcrição GênicaRESUMO
The nucleotide sequence of the simian virus 40 DNA segment that lies between the HindIII restriction endonuclease cleavage site at map position 0.324 and the AtuI cleavage site at 0.261 has been determined by the base-specific partial chemical degradation procedure of Maxam and Gilbert. This region comprises 335 base pairs and represents 6.4% of the total SV40 genome and 41% of the restriction fragment Hind-B. It connects in the clockwise direction to the restriction fragment Hind-I (described in the preceding paper). Hind-B is situated in the second half of the early transcription region of the SV40 genome and encodes information (including the termination signal) for the early protein large-T antigen. in the part of Hind-B described here, the DNA strand that has the same polarity as early 19-S mRNA defines only one open reading frame for translation; the other two are blocked by multiple triplets corresponding to termination codons. The open reading frame is part of one that runs throughout much of the early region: from the second splicing boundary of the large-T gene (position 0.534) to the information for the large-T stop signal (position 0.174) near the Hind-B/Hind-G junction, and which together with the non-contiguous DNA segment from position 0.65 (site of the information for the large-T start signal) to 0.69 (the first splicing boundary) codes for the entire large-T antigen.
Assuntos
Antígenos Virais/genética , DNA Viral , Desoxirribonucleases de Sítio Específico do Tipo II , Vírus 40 dos Símios/genética , Antígenos Virais de Tumores , Sequência de Bases , Enzimas de Restrição do DNA , Desoxirribonuclease HindIIIRESUMO
We here report the nucleotide sequence of the SV40 DNA fragment Hind C - Hap 2. The fragment was labeled at the 5'-ends by means of polynucleotide kinase and gamma-(32P) ATP and digested with a suitable restriction enzyme. The separated products were then partially degraded with the base-specific reagents dimethyl-sulphate or hydrazine followed by direct analysis on polyacrylamide gel. The Hind C - Hap 2 sequence is 126 base pairs long and one of the three possible reading frames for translation does not contain any termination codon. So, although no protein is known to be encoded by this region, the possibility cannot yet be completely ruled out. The sequence also contains several AT-rich blocks.