PubMed Health⌕ Search

SEARCH · PubMed Health

Results for “DNA, Complementary”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 757 records · Page 42Linked to original sources
PubMed1984

Cloned infectious complementary DNA of the poliovirus Sabin 1 genome: biochemical and biological properties of the recovered virus.

A complete cDNA copy of the genome of the attenuated type 1 poliovirus vaccine (Sabin 1) strain was constructed and inserted into the EcoRI site of the plasmid pBR325. When cultured mammalian cells were transfected with this recombinant plasmid, 20 to 50 poliovirus plaques per 10 micrograms plasmid DNA were observed. Fingerprints of the RNA of the recovered virus showed no changes when compared with those of the parental virus genome, an observation indicating that the primary structure of the cloned cDNA is a reflection of authentic poliovirus RNA. The recovered virus had the same properties as those of the Sabin 1 strain in regard to antigenicity, sensitivity to temperature, and dependency on bicarbonate concentration. These results suggest that the virus obtained by DNA transfection is indistinguishable from the Sabin 1 strain. The recombinant plasmid could therefore be used as a stable repository of the virus and as inoculum for the oral polio live vaccine.

Animals↗
PubMed1984

Screening recombinant phage M13 plaques with RNA probes; a one-step procedure which identifies clones containing either of the complementary DNA strands.

We describe a method for detecting specific DNA sequences cloned in M13 phage vectors, based on the procedure of Woo (in Wu, R., Methods in Enzymology, Vol. 68, Academic Press, New York, 1979, pp. 389-395). M13 plaques are adsorbed to a nitrocellulose filter that has been pre-saturated with bacteria. The filter is incubated on an agar plate to amplify the phage; the DNA is alkali-denatured and then hybridized with a radioactive RNA probe. Unlike standard procedures, this method detects and distinguishes M13 plaques containing phage particles which harbor either the coding or non-coding (RNA-like) DNA strand, when single-stranded RNA is used as probe. We have optimized this procedure with M13 clones containing mouse histidine tRNA gene sequences and have used it to determine the sequence of both strands of a mouse glycine tRNA gene.

Animals↗
PubMed1987

The nucleotide sequence of complementary DNA and the deduced amino acid sequence of peroxisomal catalase of the yeast Candida tropicalis pK233.

We report the isolation and nucleotide (nt) sequence determination of cDNA encoding peroxisomal catalase (Cat) from the yeast Candida tropicalis pK233. The catalase cDNA (Cat) has a single open reading frame (ORF) of 1455 nt, encoding a protein of 484 amino acids (aa), not including the initiator methionine. The Mr of the protein is 54767. Codon use in the gene is not random, with 90.9% of the aa specified by 25 principal codons. The principal codons used in the expression of Cat in C. tropicalis are similar to those used in the expression of the fatty acyl-CoA oxidase gene of C. tropicalis and of highly expressed genes in Saccharomyces cerevisiae. Cat shows 48.0%, 49.7%, and 48.3% aa identity with human, bovine, and rat catalases, respectively, and 44.3% aa identity with catalase T of S. cerevisiae. The 3 aa of bovine liver catalase previously postulated to participate in catalysis and 79.5% of those aa in the immediate environment of hemin, the prosthetic group of catalase, are conserved in Cat of C. tropicalis.

Amino Acid Sequence↗
PubMed1988

Complementary DNA cloning and sequencing of rat ovarian basic fibroblast growth factor and tissue distribution study of its mRNA.

Three cDNA clones encoding rat basic fibroblast growth factor (FGF) were isolated from 10(6) independent clones prepared from a pregnant mare serum gonadotropin (PMSG)-stimulated rat ovarian cDNA library. One of the cDNA clones contained the entire coding sequence for basic FGF. The other two possessed the sequence coding the carboxy terminal 61 amino acids of rat basic FGF, the putative upstream intron sequence, and a 3'-noncoding region. The cDNAs encoding rat basic FGF predict a molecule consisting of 154 amino acid residues, which is one amino acid shorter than the human and bovine basic FGF. Otherwise, there are only 5 conservative amino acid substitutions between the rat and the human/bovine sequences. Poly A+ RNA from brain cortex and hypothalamus show a single 6.0 kb band that hybridizes to the cloned cDNA probe by Northern analyses. The observation that basic FGF mRNA is below the limits of detection in adrenal, spleen, heart, lung, kidney, liver, stomach, small intestine, large intestine, testis, and ovary support the notion that the that the high levels of the protein found in these tissues is due to storage of the mitogen in the extracellular matrix and not continuous gene expression. The significance of the abundance of mRNA in tissues which are not undergoing either active angiogenesis or cell proliferation (hypothalamus and brain cortex) is unclear but emphasizes the potential neuronotrophic function of basic FGF.

Animals↗
PubMed1989

Complementary DNA for the mouse homolog of the small subunit of human cytochrome b558.

Cytochrome b558, a heterodimeric protein, is considered to be a component of the microbicidal oxidase in phagocytes. We have isolated a cDNA clone for the mouse homolog of the small subunit of human cytochrome b558. Comparison of the predicted amino acid sequence with that of the human protein reveals a high degree of homology (87%). As expected, the mRNA is expressed abundantly in peritoneal neutrophils and, to a lesser extent, in peritoneal macrophages. The transcript is also abundant in the mouse kidney. Although the spleen and small intestine contain considerable amounts of the mRNA, only trace amounts are detected in the brain, liver, testis and heart. The distribution, especially its high expression in the kidney, may suggest another function of the small subunit.

Amino Acid Sequence↗
PubMed1989

Complementary DNA structure of the high molecular weight rat insulin-like growth factor binding protein (IGF-BP3) and tissue distribution of its mRNA.

Insulin-like growth factors (IGFs) found in extracellular fluids are bound to specific binding proteins. Recently a high molecular weight IGF-binding protein (IGF-BP3) has been isolated from porcine ovarian follicular fluid based on its inhibition of follicle stimulating hormone-stimulated estradiol production in rat granulosa cells. The complete primary structure of the porcine IGF-BP3 was deduced by molecular cloning. Using the porcine cDNA as a probe, we have now isolated and characterized cDNAs encoding rat IGF-BP3 from a pregnant mare serum gonadotropin-stimulated ovarian library. The predicted amino acid sequence revealed a mature polypeptide consisting of 265 amino acids with 18 cysteines and 4 potential Asn-linked glycosylation sites. Northern analysis of the IGF-BP3 mRNA in rat tissues showed a single 2.6 kb band in liver, kidney, stomach, heart, adrenal, ovary, testis, spleen, lung, small and large intestine in varying amounts, but the message is below the limit of detection in hypothalamus and brain cortex.

Amino Acid Sequence↗
PubMed1990

Expression and cloning of complementary DNA for a human enzyme that repairs O6-methylguanine in DNA.

A cell line with an increased resistance to alkylating agents and an extremely high level of O6-methylguanine-DNA methyltransferase activity was isolated after transfection of methyltransferase-deficient Mer- cells with a cDNA library, prepared from methyltransferase-proficient human Mer+ (Raji) cells. Sodium dodecyl sulfate/polyacrylamide gel electrophoresis analysis revealed that a protein, with a molecular weight of approximately 25,000, accepted 3H label from DNA that had been treated with [3H]methylnitrosourea. Since the cDNA for methyltransferase was integrated into the chromosomal DNA, it was recovered by using the polymerase chain reaction. When the cDNA placed in an expression vector p500 was introduced into Mer- cells, the cells acquired an increased resistance to alkylating agents and exhibited a high level of O6-methylguanine-DNA methyltransferase activity. From the transformants the cDNA could be recovered as a part of the autonomously replicating plasmid. The nucleotide sequence of the cDNA was determined, and an open reading frame comprising 207 amino acid residues was found. The molecular weight of methyltransferase, calculated from the predicted amino acid sequence, was 21,700. The predicted amino acid sequence of the human methyltransferase exhibits an intensive homology with those of the bacterial counterparts, Ada and Ogt proteins of Escherichia coli and Dat protein of Bacillus subtilis, especially around possible methyl acceptor sites.

Base Sequence↗
PubMed2003

Translational machinery of channel catfish: II. Complementary DNA and expression of the complete set of 47 60S ribosomal proteins.

Ribosomal protein genes have become widely used as markers for phylogenetic studies and comparative genomics, but they have not been available in fish. We have cloned and sequenced a complete set of all 47 60S ribosomal protein cDNAs from channel catfish (Ictalurus punctatus), of which 43 included the complete protein encoding regions. Most ribosomal protein mRNAs in channel catfish are highly similar to their mammalian counterparts. However, L4, L14, and L29 are significantly shorter in channel catfish than in mammals due to deletions in the 3' end of the gene. Two distantly related L5 cDNAs, L5a and L5b, were found in channel catfish. L5a is more similar to L5 in other vertebrates, while L5b showed significant levels of divergence, suggesting independent evolution of the two L5-encoding genes. The 47 ribosomal protein genes are generally highly expressed and together account for 11-14% of overall gene expression, depending on the tissues. Expression levels were highly variable both within a single tissue among different ribosomal protein genes, and among tissues with regard to a single ribosomal protein gene. Strong tissue preference expression was also observed for some ribosomal proteins. This set of ribosomal protein gene sequences represents one of the most complete sets from any single organism and will aid in fish phylogenetic and comparative genomic studies.

Alternative Splicing↗
PubMed1997

Characterization and phylogenetic significance of rhinoceros luteinizing hormone beta (LHbeta) subunit messenger RNA structure, complementary DNA sequence and gene copy number.

The luteinizing hormone (LH) beta subunit gene is expressed in the pituitary glands of all mammals, whereas the closely related chorionic gonadotropin (CG) beta subunit genes have been identified only in primates and equids, and are expressed in placenta. In the case of horses, there is a single-copy equine (e) luteinizing hormone/chorionic gonadotropin hormone beta subunit gene (eLH/CGbeta) that (1) is expressed in both pituitary gland and placenta, (2) encodes a characteristic carboxyl terminal peptide (CTP) extension, and (3) transcribes an atypically elongated 5'-untranslated region (UTR) in both pituitary and placenta. However, it is not known whether similar expression patterns and gene locus characteristics may be exhibited by other members of the order Perissodactyla (equid, rhinoceros and tapir species). To begin to investigate these possibilities, we undertook analysis of the rhinoceros (rn or rhino) LH/(CG?)beta gene locus and the rnLHbeta cDNA. Total RNA isolated from the pituitary gland of a female white rhino was used as template for amplifying rnLHbeta cDNA by reverse transcription-polymerase chain reaction. Following cloning of the amplified cDNA, nucleotide (nt) and deduced amino acid sequences were determined. The first in-frame stop codon occurred at codon position +122, suggesting that the rnLHbeta subunit does not contain a CTP. To assess gene copy number, Southern blot analysis of Indian rhino genomic DNA was performed. The resulting simple hybridization pattern indicated that, as in the horse and donkey, there is a single-copy gene at the rnLH/(CG?)beta gene locus. Primer extension mapping of the pituitary transcriptional start site of the rnLHbeta subunit gene revealed an 8 nt 5'-UTR which is similar to that reported for the majority of mammalian LHbeta transcripts. Northern analysis was consistent with the transcriptional start site findings. We postulate from these data that rhinos diverged from equids prior to the occurrence of the mutations causing CTP expression and adoption of a non-consensus 5'-UTR/proximal promoter region. However, these findings do not rule out the possibility of expression of a placental CGbeta subunit lacking a CTP in rhinos.

Amino Acid Sequence↗
PubMed1986

Complementary DNA derived structure of the amino-terminal domain of human apolipoprotein B and size of its messenger RNA transcript.

In this paper the sequence of a 5.2-kilobase (kb) cDNA covering the amino-terminal domain of human apolipoprotein B-100 (apoB-100) is reported. The cDNA-derived protein sequence provides the primary structure of 1748 amino acids. This segment of apoB-100 is more hydrophilic than hydrophobic and contains short stretches of predicted helical and beta structures that are interrupted by beta turns. Blotting analysis of RNA isolated from fetal human and adult monkey tissues and various human cell lines showed synthesis of apoB mRNA by liver and intestine and by cells of hepatic (HepG2) and intestinal (Caco-2) origin. The isolation and characterization of overlapping cDNA clones, which provide a nearly full-length copy of human apoB-100, are also reported. From the length of these clones the size of the cytoplasmic apoB mRNA is estimated to be 14.0 kb and codes for a protein of approximately 512,000 daltons.

Adult↗
PubMed1986

Amino acid sequence of a chicken heat shock protein derived from the complementary DNA nucleotide sequence.

The complete nucleotide sequence for a chicken heat shock protein (hsp108) was determined from cDNA clones isolated from hen oviduct and bursal lymphoma recombinant DNA libraries. This protein has certain biochemical similarities to the progesterone receptor, but it is clearly distinct from it. The initial cDNA clone, isolated from a chicken oviduct cDNA library, was detected by antibody screening and hybrid-selected translation [Zarucki-Schulz, T., Kulomaa, M. S., Headon, D. R., Weigel, N. L., Baez, M., Edwards, D. P., McGuire, W. L., Schrader, W. T., & O'Malley, B. W. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 6358-6362]. The earlier clones were used to screen for additional cDNAs, and cDNAs that define the entire mRNA sequence of hsp108 have been obtained. The nucleotide sequence codes for peptides present in hsp108 as determined by protein microsequencing. The 5' end of the mRNA was determined by primer extension studies. The mRNA contains a noncoding region of 101 nucleotides upstream from the predicted initiation codon. The 3' untranslated region contains 244 nucleotides beyond the termination codon, and it contains a predicted polyadenylation signal 26 nucleotides from the end of the complete cDNA. The coding region of 2385 nucleotides corresponds to a polypeptide chain of 795 amino acids, giving a molecular weight of 91,555 for the hsp108 protein. In another paper, evidence is presented that hsp108 shows a high degree of amino acid sequence homology with two heat shock proteins, hsp90 (yeast) and hsp83 (Drosophila), and is indeed inducible by heat shock [Sargan, D. R., Tsai, M.-J., & O'Malley, B. W. (1986) Biochemistry (following paper in this issue)].

Amino Acid Sequence↗
PubMed1986

Complementary DNA sequence of lamprey fibrinogen beta chain.

The cDNA sequence of the beta chain of lamprey fibrinogen has been determined. To that end, an oligonucleotide probe was synthesized that corresponded to an amino acid sequence from the carboxy-terminal region of the lamprey fibrinogen beta chain. The insert actually began with residue 3 of the fibrin beta chain; it ran through to a terminator codon following the carboxy-terminal residue at position 443 and then continued for an additional 606 nucleotides of noncoding sequence to its 3' end. The inferred amino acid sequence was verified by comparison with assorted cyanogen bromide fragments isolated from the beta-chain protein, including two carbohydrate-containing peptides that corresponded to segments containing the carbohydrate-attachment consensus sequence. Overall, the lamprey chain is 49% identical with the beta chain from human fibrinogen. This is the same degree of resemblance as was found for the lamprey and human gamma chains. Moreover, the principal regions of conservation are the same in both the beta and gamma chains. Differences and similarities in the physiological behavior of the two fibrinogens are assessed in terms of the observed amino acid replacements.

Amino Acid Sequence↗
PubMed1987

Mammalian alpha-polymerase: cloning of partial complementary DNA and immunobinding of catalytic subunit in crude homogenate protein blots.

A new polyclonal antibody against the alpha-polymerase catalytic polypeptide was prepared by using homogeneous HeLa cell alpha-polymerase. The antibody neutralized alpha-polymerase activity and was strong and specific for the alpha-polymerase catalytic polypeptide (Mr 183,000) in Western blot analysis of crude extracts of HeLa cells. The antibody was used to screen a cDNA library of newborn rat brain poly(A+) RNA in lambda gt11. A positive phage was identified and plaque purified. This phage, designated lambda pol alpha 1.2, also was found to be positive with an antibody against Drosophila alpha-polymerase. The insert in lambda pol alpha 1.2 (1183 base pairs) contained a poly(A) sequence at the 3' terminus and a short in-phase open reading frame at the 5' terminus. A synthetic oligopeptide (eight amino acids) corresponding to the open reading frame was used to raise antiserum in rabbits. Antibody affinity purified from this serum was found to be immunoreactive against purified alpha-polymerase by enzyme-linked immunosorbent assay and was capable of immunoprecipitating alpha-polymerase. This indicated the lambda pol alpha 1.2 insert encoded an alpha-polymerase epitope and suggested that the cDNA corresponded to an alpha-polymerase mRNA. This was confirmed in hybrid selection experiments using pUC9 containing the cDNA insert and poly(A+) RNA from newborn rat brain; the insert hybridized to mRNA capable of encoding alpha-polymerase catalytic polypeptides. Northern blot analysis of rat brain poly(A+) RNA revealed that this mRNA is approximately 5.4 kilobases.

Amino Acid Sequence↗
PubMed1987

Cloning and sequence determination of a complementary DNA related to human liver microsomal cytochrome P-450 S-mephenytoin 4-hydroxylase.

A cDNA sequence related to the human cytochrome P-450 responsible for S-mephenytoin 4-hydroxylation (P-450MP) has been isolated from a human liver bacteriophage lambda gt11 library with antibodies specific for P-450MP. The total length of the cDNA is 2.5 kilobases (kb), of which there is a 1.6-kb EcoRI fragment coding for all but five amino acids corresponding to the N-terminus of the protein and including a small noncoding region at the 3' end. This 1.6-kb fragment has been sequenced and used as a probe to analyze human genomic DNA and liver RNA. The sequence shows extensive sequence similarity with that of rabbit liver cytochrome P-450 progesterone 21-hydroxylase [Tukey, R. H., Okino, S., Barnes, H., Griffin, K. J., & Johnson, E. F. (1985) J. Biol. Chem. 260, 13347-13354], and this cDNA, like the rabbit clone, appears to be part of a multigene family. At least two liver mRNA species, 2.2 kb and 3.5 kb, hybridize to the cDNA sequence. The cloning of this gene should aid in analyzing the molecular basis for the genetic polymorphism of S-mephenytoin 4-hydroxylation reported in humans.

Amino Acid Sequence↗
PubMed1987

Molecular cloning and sequence analysis of complementary DNA encoding rat mammary gland medium-chain S-acyl fatty acid synthetase thio ester hydrolase.

Poly(A)+ RNA from pregnant rat mammary glands was size-fractionated by sucrose gradient centrifugation, and fractions enriched in medium-chain S-acyl fatty acid synthetase thio ester hydrolase (MCH) were identified by in vitro translation and immunoprecipitation. A cDNA library was constructed, in pBR322, from enriched poly(A)+ RNA and screened with two oligonucleotide probes deduced from rat MCH amino acid sequence data. Cross-hybridizing clones were isolated and found to contain cDNA inserts ranging from approximately 1100 to 1550 base pairs (bp). A 1550-bp cDNA insert, from clone 43H09, was confirmed to encode MCH by hybrid-select translation/immunoprecipitation studies and by comparison of the amino acid sequence deduced from the DNA sequence of the clone to the amino acid sequence of the MCH peptides. Northern blot analysis revealed the size of the MCH mRNA to be 1500 nucleotides, and it is therefore concluded that the 1550-bp insert (including G X C tails) of clone 43H09 represents a full- or near-full-length copy of the MCH gene. The rat MCH sequence is the first reported sequence of a thioesterase from a mammalian source, but comparison of the deduced amino acid sequences of MCH and the recently published mallard duck medium-chain S-acyl fatty acid synthetase thioesterase reveals significant homology. In particular, a seven amino acid sequence containing the proposed active serine of the duck thioesterase is found to be perfectly conserved in rat MCH.

Amino Acid Sequence↗
PubMed1987

Repeating structure of chick tropoelastin revealed by complementary DNA cloning.

A cDNA library was constructed from chick aorta poly(adenylic acid)-containing RNA in the expression vector pEX1. Several clones were identified by screening the library with a polyclonal antiserum raised against chick tropoelastin and confirmed by DNA sequencing. Analysis of the deduced amino acid sequence, corresponding to the mature tropoelastin and most of the signal peptide, revealed that the molecule is composed of at least 8, and possibly 13, repeating units. The common features of each unit include an N-terminal region composed largely of alanines and lysines and ending with an aromatic amino acid, followed by a GAG span and then a C-terminal region consisting mostly of valines, prolines, and glycines often present in several copies of the sequence (VPGV). This structure is discussed in terms of the functional properties of the molecule.

Amino Acid Sequence↗