RESUMO
A cDNA library of the bovine mammary gland constructed in pBR322 was screened by mRNA hybrid-selected translation and by differential hybridization. Several immunoglobulin (Ig) lambda light-chain clones were identified and sequenced. Nucleotide sequence comparison of bovine and human Ig lambda chains showed a high degree of homology for constant regions and for J regions. The amino acid (aa) sequence encoded by the constant region of the bovine Ig lambda chain cDNA contains 107 aa with differences at 24 aa positions from the human Ig lambda chain. Three complementarity-determining regions (CDR1,2,3) characteristic of the variable region of bovine Ig lambda chain cDNA can be distinguished. The bovine and human sequences display good homology in the framework region 3 (FR3) but only patches of homology throughout the FR2 region. The 5' end of the bovine Ig lambda chain cDNA fragment of clone 1-14E contains five stop codons: two in CDR1, one in FR1 and two in the hydrophobic prepeptide region. These data suggest that the Ig lambda mRNA of clone 1-14E is transcribed from the V lambda pseudogene.
Assuntos
Clonagem Molecular , DNA/genética , Genes de Imunoglobulinas , Cadeias lambda de Imunoglobulina/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Bovinos , Enzimas de Restrição do DNA , Feminino , Glândulas Mamárias Animais/imunologia , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Plasmídeos , Biossíntese de Proteínas , RNA Mensageiro/genéticaRESUMO
Genes encoding luminescence of Photobacterium leiognathi have been cloned in Escherichia coli. The luminescent clones were readily apparent. Among them, a clone containing a recombinant plasmid with a 13.5-kb insertion was identified. This DNA fragment contained all of the luminescence-encoding genes. The luciferase-encoding genes (lux) in this DNA fragment were localized. We have sequenced a part of the cloned lux region and identified the luxA, luxB and luxG genes encoding the alpha and beta subunits of luciferase and a gamma protein with an Mr of 26,180, respectively. The analysis of deduced amino acid sequences and comparison with known luciferase sequences from Vibrio harveyi, indicate the common origin of these proteins.
Assuntos
Genes Bacterianos , Luciferases/genética , Medições Luminescentes , Photobacterium/genética , Aciltransferases/genética , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Sequência de Bases , Clonagem Molecular , DNA Bacteriano/genética , Dados de Sequência Molecular , Óperon , Mapeamento por RestriçãoRESUMO
The initiation sites of the Galleria mellonella L. nuclear polyhedrosis virus (G.m. NPV) DNA replication were revealed. For this purpose SCLd 135 cells permitting the G.m. NPV productive reproduction were transformed by the recombinant plasmids containing the viral genome individual fragments in pRSF 2124 and pBR 322 vectors. It was revealed that 2 of the 32 recombinant plasmids can autonomously replicate in the eucaryotic cells. According to the Maxam-Gilbert method the DNA G.m. NPV fragment (1300 bp) primary structure of pHBR plasmid was determined. The structure analysis revealed the typical regulator signals as in the replicons. The possible regulation mechanism of the DNA G.m. NPV synthesis initiation was supposed.
Assuntos
Replicação do DNA , Vírus de Insetos/genética , Sequência de Bases , Replicação ViralRESUMO
The nucleotide sequence coding for human angiogenin has been deduced from the published amino acid sequence with the use of codons preferentially utilized in highly expressed E. coli genes. It was divided into forty-three oligonucleotides, which were synthesized by automatic gene assembler and then joined by DNA ligase into three double-stranded blocks, the blocks were consequently cloned and ligated in M13mp8 phage, and the resultant 389-bp DNA sequence coding for human angiogenin was analysed by chain-terminator sequencing technique.
Assuntos
Indutores da Angiogênese/genética , Clonagem Molecular , Genes Sintéticos , Genes , Substâncias de Crescimento/genética , Proteínas de Neoplasias/genética , Ribonuclease Pancreático , Sequência de Aminoácidos , Sequência de Bases , Códon , Colífagos/genética , Vetores Genéticos , Humanos , Dados de Sequência MolecularRESUMO
Full-length DNA copies of M-gene of remantadine-sensitive and remantadine-resistant variants of the influenza virus strain A/FPV/Weybridge (H7N7) have been synthesised and cloned. Complete nucleotide sequences of both cDNAs were determined by the Maxam-Gilbert method. There are three nucleotide substitutions, two of which lead to amino acid changes in M1 and M2 proteins. The existence of M3 protein, a polypeptide 68 amino acids long, encoded by the negative strand of RNA, is suggested. Amino acid changes in M2 and M3 proteins and their relation to the remantadine resistance are discussed.
Assuntos
Adamantano/análogos & derivados , Genes Virais , Vírus da Influenza A Subtipo H7N7 , Vírus da Influenza A/genética , Rimantadina/farmacologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Embrião de Galinha , DNA/genética , DNA Viral/genética , Resistência Microbiana a Medicamentos , Vírus da Influenza A/efeitos dos fármacos , Dados de Sequência Molecular , Proteínas Virais/genéticaRESUMO
The poly(A)-containing mRNA from human pituitary and prolactinoma have been purified and translated in the cell-free system from rabbit reticulocytes. mRNA from prolactinoma was shown to be enriched with specific prolactin mRNA. DNA complementary to the prolactin mRNA from human pituitary was obtained and cloned. Sequencing of the 900 bp insert by the Maxam-Gilbert technique suggested the cDNA cloned to cole for the previously published amino acid sequence, mismatches with mRNA from prolactinoma occurring at the third positions of codons and thus not causing amino acid substitutions.
Assuntos
Clonagem Molecular , DNA/genética , Hipófise/análise , Prolactina/genética , RNA Mensageiro/genética , Sequência de Bases , DNA/análise , DNA/síntese química , Eletroforese em Gel de Poliacrilamida , Humanos , Dados de Sequência Molecular , Prolactina/análise , RNA Mensageiro/análiseRESUMO
cDNA coding for the human pro-opiomelanocortin (POMC) has been cloned and sequenced. It codes for full size amino acid sequence of POMC and furthermore, contains most part of the 3'-terminal noncoding mRNA region and 60 nucleotides coding for signal peptide.
Assuntos
Clonagem Molecular , DNA/genética , Hipófise/análise , Pró-Opiomelanocortina/genética , RNA Mensageiro/genética , Sequência de Bases , DNA/biossíntese , Humanos , Pró-Opiomelanocortina/isolamento & purificaçãoRESUMO
Total RNA was extracted from Mustella vison pituitary gland, and cDNA for proopiomelanocortin mRNA was synthesised and cloned. A 600 b. p. insert encoding for total ACTH, beta LPH and 3'-nontranslated end of the mRNA was sequenced using the Maxam-Gilbert technique.
Assuntos
Clonagem Molecular , DNA/biossíntese , Hipófise/análise , Pró-Opiomelanocortina/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA/genética , Vison , Dados de Sequência Molecular , Plasmídeos , RNA Mensageiro/genéticaRESUMO
Nucleotide sequence of the Photobacterium leiognathi DNA containing genes of alpha and beta subunits of luciferase has been determined. We also deduced amino acid sequence and molecular mass of luciferase and localized luciferase genes in the sequenced DNA fragment.
Assuntos
Genes Bacterianos , Luciferases/genética , Photobacterium/genética , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , Dados de Sequência Molecular , Photobacterium/enzimologiaRESUMO
Complete nucleotide sequence of the cloned full-length DNA copy of the influenza virus A (H1N1) neuraminidase gene has been determined. The predicted amino acid sequence is compared with sequences of neuraminidases from other influenza virus strains. A section of the neuraminidase is found to be homologous to the chicken lysozyme catalytic centre.
Assuntos
DNA Viral , Genes Virais , Vírus da Influenza A Subtipo H1N1 , Vírus da Influenza A/genética , Neuraminidase/genética , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , DNA Viral/biossíntese , Vírus da Influenza A/enzimologia , Sequências Repetitivas de Ácido NucleicoRESUMO
Complete nucleotide sequence of the cloned full-length DNA copy of the influenza virus A (H1N1) RNA segment 5 has been determined. The degree of nucleotide difference between two variants of A/PR/8/34 strain is estimated. The possible secondary structure of the segment 5 is deduced from the nucleotide sequence of some clones.
Assuntos
DNA Viral , Genes Virais , Vírus da Influenza A/genética , Nucleoproteínas , Proteínas do Core Viral , Proteínas Virais/genética , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , DNA Viral/biossíntese , DNA Viral/genética , Proteínas do Nucleocapsídeo , Sequências Repetitivas de Ácido NucleicoRESUMO
Complete nucleotide sequence of the cloned full-length DNA copy of the influenza virus A (H1N1) RNA segment 8 has been determined. A section of the hypothetical protein coded for by the negative strand of the segment 8 is found to be homologous to the trypsin catalytic centre.
Assuntos
DNA Viral , Genes Virais , Vírus da Influenza A/genética , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , DNA Viral/biossíntese , Sequências Repetitivas de Ácido Nucleico , Proteínas Virais/genéticaRESUMO
The double-stranded DNA copy of the matrix protein (M) gene of the influenza virus A/USSR/90/77 (H1N1) has been inserted in PstI site of plasmid pBR322 and cloned in E. coli. The full-length DNA copy of the M gene has been sequenced using Maxam-Gilbert method. Analysis of nucleotide sequence of segment 7 RNA of influenza virus A/USSR/90/77 and nucleotide substitutions, as compared with known primary structures of segment 7 RNA for other strains, is presented. A hypothetical model of secondary structure of segment 7 RNA of influenza virus and repeating sequences at nucleotide and amino acid levels, revealed in the central region of M1 protein, are discussed.
Assuntos
Vírus da Influenza A Subtipo H1N1 , Vírus da Influenza A/análise , RNA Viral/análise , Sequência de Aminoácidos , Sequência de Bases , Vírus da Influenza A/genética , Conformação de Ácido Nucleico , RNA Viral/genética , Sequências Repetitivas de Ácido Nucleico , Proteínas Virais/análise , Proteínas Virais/biossínteseRESUMO
The complete nucleotide sequence of the cloned full-length DNA copy of A/Kiev/59/79 (H1N1) influenza virus nucleoprotein gene has been determined. This strain is shown to be the natural recombinant that inherited its nucleoprotein gene from contemporary H3N2-influenza strains. The comparison with other NP-genes reveals the probable localization of antigenic determinants and phosphorylation site of the NP-protein.
Assuntos
DNA Viral/análise , Genes Virais , Vírus da Influenza A Subtipo H1N1 , Vírus da Influenza A/genética , Nucleoproteínas , Proteínas do Core Viral/genética , Sequência de Aminoácidos , Sequência de Bases , DNA Viral/genética , Humanos , Vírus da Influenza A/classificação , Proteínas do Nucleocapsídeo , Recombinação Genética , Sorotipagem , Proteínas do Core Viral/análiseRESUMO
The complete nucleotide sequence of the cloned full-length DNA copy of A/Kiev/59/79 (H1N1) influenza virus hemagglutinin gene has been determined. The comparison with the other hemagglutinin structures reveals the divarication of evolutionary pathway of the H1N1-influenza viruses.
Assuntos
DNA Viral/análise , Genes Virais , Hemaglutininas Virais/genética , Vírus da Influenza A Subtipo H1N1 , Vírus da Influenza A/genética , Sequência de Aminoácidos , Sequência de Bases , DNA Viral/genética , Hemaglutininas Virais/análise , Humanos , Vírus da Influenza A/classificação , Filogenia , SorotipagemRESUMO
Reverse transcription produced DNA-copies of the 3'-terminal sequence of mouse encephalomyocarditis virus RNA. Cloning and sequencing of this cDNA afforded the structure of the 3'-terminus of viral RNA over the length of 2988 nucleotide residues. The probable sites of proteolytic cleavage of the immature protein have been determined and the homology with the respective poliovirus proteins has been established.
Assuntos
Vírus da Encefalomiocardite/genética , RNA Viral/genética , Sequência de Aminoácidos , Sequência de Bases , DNA/genética , Hibridização de Ácido Nucleico , Proteínas ViraisRESUMO
The complete nucleotide sequence of the cloned full-length DNA copy of the influenza virus A/Kiev/59/79 (H1N1) neuraminidase gene has been determined. The comparison with the other neuraminidases reveals the differences in localization of antigenic determinants between N1 and N2 subtypes and divarication of evolutionary pathways of the modern H1N1-influenza viruses.
Assuntos
DNA Viral/análise , Genes Virais , Vírus da Influenza A Subtipo H1N1 , Vírus da Influenza A/genética , Neuraminidase/genética , Sequência de Bases , Humanos , Vírus da Influenza A/enzimologiaRESUMO
Three fragments of a DNA copy complementary to tre hepatitis A virus RNA have been isolated from the pHAV-VPI-22 plasmid and cloned in M 13 mp 8 vector. The 140, 250 and 390 b. p. fragments corresponded to the viral genome fragment coding for VP1 protein. Single-stranded DNAs of the hybrid phages with 250 b. p. and 390 b. p. inserts have been used for radioactive probe synthesis. The specificity of the probes for viral RNA was confirmed in hybridization with hepatitis A virus RNA purified from the virus-infected cell culture. The hybridization was shown to detect up to 10(-12)-10(-13) g of the virus. A principal possibility to use the probes in diagnostic purposes was demonstrated by the virus detection in blood samples obtained from patients with hepatitis A infection.
Assuntos
Colífagos/genética , Sondas de DNA , Hepatovirus/genética , Hibridização de Ácido Nucleico , RNA Viral/genética , DNA Viral/genética , PlasmídeosRESUMO
A scheme for evolutionary interrelations of the H1-subunits of influenza hemagglutinin genes is proposed for the natural variants of influenza A virus of the H1N1-subtype. It is based on experimental data obtained by the authors and those reported in the literature. Differences among these viral isolates in their amino acid sequences and in the reaction of hemagglutinin inhibition obtained with a set of monoclonal antibodies are compared. The distinctions in the ability of the viruses to react with several monoclonal antibodies are attributed to differences in the primary structures of their hemagglutinins. Some aspects of hemagglutinin gene evolution are discussed in relation to vaccination.
Assuntos
Genes Virais , Hemaglutininas Virais/genética , Vírus da Influenza A Subtipo H1N1 , Vírus da Influenza A/genética , Sequência de Aminoácidos , Evolução BiológicaRESUMO
An efficient method to obtain the mutant genes for human leucocyte alpha 2-interferon (IFN) has been elaborated. The technique includes the following main stages: cloning of interferon gene in M13mp8 DNA; isolation of double-stranded hybrid DNA complex, containing IFN gene as a single-stranded fragment; selective modification of a single-stranded hybrid DNA by sodium bisulphite; the repair of hybrid DNA by DNA polymerase I from Escherichia coli, transformation of Escherichia coli JN103 cells by double-stranded circular DNA, containing the selectively modified gene IFN. The technique is based on the protection of bacteriophage M13 genome from mutagen induced damage by means of converting phage DNA into the double-stranded structure leaving the single-stranded fragment to be mutagenized prone to mutagen action. This is achieved by reannealing of single-stranded M13mpB DNA hydrolyzed by restriction endonuclease BamHI. The technique preserves the infectiousness of vector DNA under the conditions permitting modification of up to 10% cytosine residues in IFN gene. Every clone resulting from transformation of Escherichia coli by modified DNA carried mutations in IFN gene, identified by sequencing after Sanger.