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At least 109 records · Page 6Linked to original sources
PubMed1991

Molecular cloning and nucleotide sequence of complementary DNA encoding rabbit alpha 1-acid glycoprotein.

Alpha 1-acid glycoprotein (AGP) complementary DNA clone has been isolated from an acute phase rabbit liver library. Complete and full-length nucleotide sequence of this cDNA has been determined. The cDNA contains 35 nucleotides 5' untranslated region followed by 606 nucleotides of coding region and a 112 nucleotides 3' untranslated region. Comparison of this sequence with that of human, rat and mouse reveals a high degree of homology with human alpha 1-AGP. Northern blot analysis of messenger RNA for alpha 1-AGP in normal and acute livers demonstrates remarkable induction of transcription of this gene in rabbit in response to acute inflammation by the administration of turpentine. Such a response of AGP gene expression makes it a major member of the acute phase responsive genes in rabbit liver.

Amino Acid Sequence↗
PubMed1991

Complementary DNA cloning and nucleotide sequence of rabbit serum amyloid A protein.

A complementary DNA clone encoding serum amyloid A protein has been isolated from an acute rabbit liver cDNA library. Complete nucleotide sequence analysis reveals that the cloned gene contains a 24 bases 5' untranslated region, 369 bases coding region and a 106 bases 3' untranslated region. Primer extension analysis indicates that the full-length 5' untranslated region contains 80 nucleotides. Northern blot analysis of mRNA from normal and acute rabbit livers demonstrates that this gene is expressed constitutively at a low level and undergoes induction of transcription in response to acute inflammation by the administration of turpentine.

Amino Acid Sequence↗
PubMed1985

On the mechanism of renaturation of complementary DNA strands by the recA protein of Escherichia coli.

The renaturation of complementary DNA strands by the recA protein of Escherichia coli has been found to exhibit the following features. (i) Optimal renaturation occurs at recA protein levels below that required to saturate the DNA strands; saturating amounts of recA protein significantly reduce the rate of reaction. (ii) The reaction proceeds in the absence of a nucleotide cofactor but is markedly stimulated by ATP in the presence of 10 mM Mg2+. A similar stimulation occurs in the absence of ATP when the Mg2+ concentration is increased from 10 mM to 30-40 mM. (iii) Both the ATP-stimulated and the Mg2+-stimulated reactions follow apparent first-order kinetics. These results, taken together with the known effects of ATP and Mg2+ on the state of aggregation of recA protein, suggest that the association of recA monomers may play an important role in recA protein-promoted DNA renaturation.

Adenosine Triphosphate↗
PubMed1989

Complementary DNA coding click beetle luciferases can elicit bioluminescence of different colors.

Eleven complementary DNA (cDNA) clones were generated from messenger RNA isolated from abdominal light organs of the bioluminescent click beetle, Pyrophorus plagiophthalamus. When expressed in Escherichia coli, these clones can elicit bioluminescence that is readily visible. The clones code for luciferases of four types, distinguished by the colors of bioluminescence they catalyze: green (546 nanometers), yellow-green (560 nanometers), yellow (578 nanometers), and orange (593 nanometers). The amino acid sequences of the different luciferases are 95 to 99 percent identical with each other, but are only 48 percent identical with the sequence of firefly luciferase (Photinus pyralis). Because of the different colors, these clones may be useful in experiments in which multiple reporter genes are needed.

Adenosine Triphosphate↗
PubMed1985

Molecular cloning of a complementary DNA encoding human macrophage-specific colony-stimulating factor (CSF-1).

Complementary DNA (cDNA) clones encoding human macrophage-specific specific colony-stimulating factor (CSF-1) were isolated. One cDNA clone codes for a mature polypeptide of 224 amino acids and a putative leader of 32 amino acids. This cDNA, which was cloned in the Okayama-Berg expression vector, specifies the synthesis of biologically active CSF-1 in COS cells, as determined by a specific radioreceptor assay, macrophage bone marrow colony formation, and antibody neutralization. Most of the cDNA isolates contain part of an intron sequence that changes the reading frame, resulting in an abrupt termination of translation; these cDNA's were inactive in COS cells. The CSF-1 appears to be encoded by a single-copy gene, but its expression results in the synthesis of several messenger RNA species, ranging in size from about 1.5 to 4.5 kilobases.

Amino Acid Sequence↗
PubMed1991

Isolation and characterization of a complementary DNA (PD-1) differentially expressed by human pancreatic ductal cell tumors.

A complementary DNA (cDNA) library from a cloned subline (CD-11) of a well differentiated human pancreatic tumor cell line, HPAF, was subjected to differential screening using single stranded cDNA probes synthesized from mRNA of the well differentiated cell clone CD-11 and a poorly differentiated pancreatic tumor cell line, Panc 1. A cDNA clone (PD-1) was identified which had an insert of 626 base pairs (bp). PD-1 cDNA hybridized to a transcript of about 650 bp on Northern blot analysis, suggesting that the cDNA was close to full length. Densitometric analysis of Northern blots showed that a well differentiated pancreatic tumor line had a 5-fold higher PD-1 expression as compared to the poorly differentiated line, Panc 1. Nucleotide sequence analysis of the PD-1 cDNA and its deduced amino acid sequence showed an open reading frame of 399 bp. In addition to the open reading frame, the sequence had a 5' untranslated region of 61 bp and a 3' untranslated tail of 147 bp. The nucleotide sequence did not show any significant homology to any other sequence in the GENBANK or EMBL databases; however, the translated protein showed 35% homology to bacterial ribosomal proteins over 112 amino acids. Sequence analysis of the PD-1 cDNA and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of its in vitro transcription/translation product suggest that this gene encoded a protein of 16,000 daltons.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1988

Molecular cloning of human and rat complementary DNA encoding androgen receptors.

Complementary DNAs (cDNAs) encoding androgen receptors were obtained from human testis and rat ventral prostate cDNA libraries. The amino acid sequence deduced from the nucleotide sequences of the cDNAs indicated the presence of a cysteine-rich DNA-binding domain that is highly conserved in all steroid receptors. The human cDNA was transcribed and the RNA product was translated in cell-free systems to yield a 76-kilodalton protein. The protein was immunoprecipitable by human autoimmune antibodies to the androgen receptor. The protein bound androgens specifically and with high affinity.

Amino Acid Sequence↗
PubMed1987

Expression of functional cell-cell channels from cloned rat liver gap junction complementary DNA.

An oocyte expression system was used to test the relation between a complementary DNA (cDNA) clone encoding the liver gap junction protein and cell-cell channels. Total liver polyadenylated messenger RNA injected into oocytes induced cell-cell channels between paired oocytes. This induction was blocked by simultaneous injection of antisense RNA transcribed from the gap junction cDNA. Messenger RNA selected by hybridization to the cDNA clone and translated in oocyte pairs yielded a higher junctional conductance than unselected liver messenger RNA. Cell-cell channels between oocytes were also formed when the cloned cDNA was expressed under the control of a heat-shock promoter. A concentration-dependent induction of channels was observed in response to injection with in vitro transcribed gap junction messenger RNA. Thus, the liver gap junction cDNA encodes a protein that is essential for the formation of functional cell-cell channels.

Animals↗
PubMed1991

A new growth-regulated complementary DNA with the sequence of a putative trans-activating factor.

A new complementary DNA (cDNA) clone has been isolated by differential screening of a cDNA library. The cognate RNA of this clone, called SC1, is growth regulated in human, mouse, and hamster cell lines. Its kinetics of growth regulation (time of increase in mRNA levels, sensitivity to cycloheximide, behavior in G1-specific temperature-sensitive mutants) classify the SC1 gene as a late growth-regulated gene, like the histone genes and the genes coding for the proteins of the DNA synthesis apparatus. By run-on assay, there is a modest increase in transcriptional rates after serum stimulation, which is not sufficient to explain the sharp increase in mRNA levels. The SC1 gene localizes to human chromosome 6p21-22. In bacteria, the SC1 cDNA clone makes a protein of Mr 39,000, in agreement with the putative reading frame. The amino acid sequence derived from the cDNA sequence indicates a previously unknown gene with a domain strongly suggestive of a trans-activating domain. The SC1 gene can be considered as coding for a possible new trans-activating factor that could play an important role in the transcription of genes required for the later stages of cell cycle progression.

Amino Acid Sequence↗
PubMed1977

Human beta-globin messenger RNA. III. Nucleotide sequences derived from complementary DNA.

Sequences of human beta-globin mRNA were determined by analysis of complementary DNA. beta-mRNA was transcribed into double-stranded cDNA by RNA-dependent DNA polymerase. cDNA was cut by restriction endonucleases and the fragments were terminally labeled by means of polynucleotide kinase and [gamma-32P]ATP. After purification, fragments were degraded by snake venom phosphodiesterase. Alternatively single-stranded [32P]cDNA was prepared by transcription in the presence of [alpha-32P]dCTP and actinomycin D; the product was digested by endonuclease IV and degraded by snake venom phosphodiesterase. cDNA tracts obtained by both labeling methods enabled us to construct a sequence for the translated and 3'-terminal untranslated regions of human beta-mRNA.

Anemia, Sickle Cell↗
PubMed1979

Androgenic regulation of messenger RNA sequence complexity in accessory sexual tissues of the male rat studied with fractionated complementary DNA.

Effects of androgens on mRNA sequence complexity in the rat seminal vesicle have been investigated using complementary DNA fractionated on the basis of sequence abundance. Total cDNA complementary to poly(A)-rich RNA from normal rats was hybridised with an excess of the same RNA to controlled rot values and then the free cDNA was separated from cDNA . RNA hybrids by hydroxyapatite chromatography. Three cDNA fractions were obtained with very different hybridisation characteristics. Abundant cDNA hybridised to an excess of its parental RNA with an rot 1/2 of 2.46 x 10(-3) mol 1(-1) s and is complementary to about six or seven average-sized sequences. Use of hybrid-arrested translation in a cell-free protein-synthesising system has shown that this class of mRNA includes mRNAs coding for major androgen-dependent secretory proteins. Moderate and scarce cDNA fractions each showed more complex hybridization kinetics; computer analysis suggested each is complementary to two groups of average-sized sequences. Each cDNA fraction was hybridised to excess poly(A)-rich RNA from normal or castrated rats and the kinetics compared. Castration had no effect on the total number of sequences present in any class and did not alter the relative concentration of the scarce sequences. A small (threefold) decrease was seen in the concentration of abundant sequences with a larger (tenfold) decrease in the moderate class. Both de-reases were reversed by testosterone in vivo. The results are consistent with earlier studies where the effects of testosterone on seminal vesicle mRNA were followed using a translation assay and confirm that no gross differential effects are exerted on abundant mRNA coding for major secretory proteins. The cDNA fractions were also used to investigate the overlap in genetic expression between seminal vesicle and ventral prostate. Both tissues share all the scarce sequences in the same relative abundance. Less than 0.0015% and 0.004% of prostatic mRNA is complementary to seminal vesicle abundant and moderate sequences respectively. Similarly prostatic abundant sequences account for less than 0.004% of seminal vesicle mRNA.

Animals↗
PubMed1977

The use of a complementary DNA probe to detect accumulation of mengo RNA in infected cells pretreated with interferon.

Complementary DNA (cDNA) from Mengo virus RNA has been synthesized and used as a probe to measure the synthesis and accumulation of viral RNA in Mengo infected L cell cultures, treated or untreated with interferon. Under experimental conditions used (200 units interferon/ml and 50 virus plaque-forming units/cell) results show that there is some synthesis of Mengo virus RNA in cells treated with interferon. One hour after infection, treated cells contain three times less viral RNA than untreated cells; five hours after infection, this difference has increased to ten fold. As in the control, no fragmented Mengo virus RNA molecules were found in interferon treated cells. The smaller recovery of infectious particles from interferon treated cells as compared to RNA accumulation suggests that not only RNA accumulation is inhibited but also a step posterior in viral maturation.

Cytoplasm↗
PubMed1977

Multihormonal induction of hepatic alpha2u-globulin mRNA as measured by hybridization to complementary DNA.

A procedure is presented for the preparation of a (3)H-labeled complementary DNA (cDNA) specific for the mRNA coding for alpha(2u)-globulin, a male rat liver protein under multihormonal control that represents approximately 1% of hepatic protein synthesis. Rat liver polysomes are incubated with monospecific rabbit antiserum to alpha(2u)-globulin, which binds to the nascent alpha(2u)-globulin chains on the polysomes. These antibody-polysome complexes are then adsorbed to goat antiserum to rabbit IgG that is covalently linked to p-aminobenzylcellulose. mRNA preparations are thus obtained that contain 30-40% alpha(2u)-globulin mRNA. A labeled cDNA is made to this alpha(2u)-globulin-enriched mRNA preparation by using RNA-dependent DNA polymerase (reverse transcriptase). To remove the non-alpha(2u)-globulin sequences, this cDNA preparation is hybridized to an RNA concentration x incubation time (R(0)t) of 1000 mol of ribonucleotide per liter x sec with female rat liver mRNA, which, though it shares the vast majority of mRNA sequences with male liver, contains no alpha(2u)-globulin mRNA sequences. The cDNA remaining single-stranded is isolated by hydroxylapatite chromatography and is shown to be specific for alpha(2u)-globulin mRNA by several criteria. Good correlation was found in all endocrine states studied between the hepatic level of alpha(2u)-globulin, the level of functional alpha(2u)-globulin mRNA as assayed in a wheat germ cell-free translational system, and the level of alpha(2u)-globulin mRNA sequences as measured by hybridization to the alpha(2u)-globulin cDNA. Thus, the hormonal control of hepatic alpha(2u)-globulin synthesis by sex steroids and thyroid hormone occurs through modulation of the cellular level of alpha(2u)-globulin mRNA sequences, presumably by hormonal control of transcriptive synthesis.

Alpha-Globulins↗
PubMed1988

New BK virus episomal vector for complementary DNA expression in human cells.

The properties of pRP-c, a new vector for complementary DNA (cDNA) expression, are described. The vector contains the early region and replication origin of BK virus (BKV), a human papovavirus. Due to the presence of these BKV sequences, pRP-c replicates in human cells allowing amplification of inserted cDNAs. The promoter, intron and polyadenylation region for cDNA expression are separated by unique restriction sites and can therefore be individually excised and substituted with different transcription signals. Coding sequences of the bacterial genes for chloramphenicol-acetyl transferase (CAT) or neomycin phosphotransferase (neo) were inserted into the cDNA cloning site of pRP-c and expressed in human cells in transient assays or stable clones. In both cases expression of the inserted sequences was significantly more efficient than by using the integration vectors pSV2CAT and pSV2neo, demonstrating the advantages of episomal expression vectors in human cells. Possible uses of pRP-c to express viral and cellular cDNAs in human cells are discussed.

BK Virus↗
PubMed1987

Replication origin (oric) on the complementary DNA strand of Escherichia coli phage G4: biological properties of mutants.

Phage G4 origin of complementary DNA strand synthesis (oric) consists of three stable secondary loop structures. In a cloned 274-bp DNA fragment that is active as an ori in the filamentous phage cloning vector R199, insertion mutants have been constructed by introducing EcoRI and HindIII linkers at the base of loop III. The in vivo activity of these oric mutants (conversion of single-strand form to replicative form in the presence of rifampicin) was significantly reduced (50-70%) but not completely abolished. Nucleotide sequences and/or potential secondary structure of loop III centered at the AvaII site are therefore an important functional part of oric.

Base Sequence↗
PubMed1990

Isolation of complementary DNA clones encoding pathogenesis-related proteins P and Q, two acidic chitinases from tobacco.

Complementary DNA clones encoding two isoforms of the acidic endochitinase (chitinase, EC 3.2.1.14) from tobacco were isolated. Comparison of amino acid sequences deduced from the cDNA clones and the sequence of peptides derived from purified proteins show that these clones encode the pathogenesis-related proteins PR-P and PR-Q. The cDNA inserts were not homologous to either the bacterial form of chitinase or the form from cucumber but shared significant homology to the basic form of chitinase from tobacco and bean. The acidic isoforms of tobacco chitinase did not contain the amino-terminal, cysteine-rich "hevein" domain found in the basic isoforms, indicating that this domain, which binds chitin, is not essential for chitinolytic activity. The accumulation of mRNA for the pathogenesis-related proteins PR-1, PR-R, PR-P, and PR-Q in Xanthi.nc tobacco leaves following infection with tobacco mosaic virus was measured by primer extension. The results indicate that the induction of these proteins during the local necrotic lesion response to the virus is coordinated at the mRNA level.

Amino Acid Sequence↗
PubMed1992

Molecular cloning of rat prostate transglutaminase complementary DNA. The major androgen-regulated protein DP1 of rat dorsal prostate and coagulating gland.

Complementary DNA (cDNA) that codes for a major androgen-dependent secretory protein of rat coagulating gland and dorsal prostate, dorsal protein 1 (DP1), was isolated by molecular cloning. Recombinant DP1 cDNA clones were identified from a bacteriophage lambda gt11 rat coagulating gland expression library using an affinity purified polyclonal antibody. Amino acid sequence deduced from DNA contained sequences identical with several DP1 cyanogen bromide cleavage fragments. Northern blot hybridization of poly(A) RNA isolated from intact rat dorsal prostate and coagulating gland revealed a predominant messenger RNA (mRNA) species of approximately 3200 nucleotides. Tissue-specific expression of DP1 mRNA was indicated by the absence of DP1 mRNA in ventral prostate and other tissues of the rat. Expression of DP1 mRNA was androgen-dependent, decreasing approximately 80% 7 days after castration and increasing rapidly following androgen replacement. Southern blot analysis of restriction enzyme-digested rat DNA indicated that DP1 is encoded by a single gene and that no major genomic rearrangements accounted for its lack of expression in the dorsal prostate-derived rat Dunning tumor. Sequence comparisons revealed that rat prostate DP1 shares sequence identity with Factor XIIIa and tissue transglutaminase, including the active center, GQCWVF, indicating that DP1 is a member of the transglutaminase gene family.

Amino Acid Sequence↗