Showing papers on "Complementary DNA published in 1980"
TL;DR: Bacterial clones containing inserted DNA sequences specific for α- Tubulin, β-tubulin,β-Actin and γ-actin have been constructed from mRNA of embryonic chick brain and are able to hybridize under stringent conditions to DNA of all vertebrates tested, as well as to sea urchin DNA, but not to yeast DNA.
1,650 citations
TL;DR: A human leukocyte interferon cDNA was enzymatically synthesized, inserted into the vector pBR322, and cloned in Escherichia coli, and protected squirrel monkeys from lethal encephalomyocarditis virus infection.
Abstract: A human leukocyte interferon cDNA was enzymatically synthesized, inserted into the vector pBR322, and cloned in Escherichia coli. The DNA sequence codes for a 23-amino acid signal peptide followed by an interferon polypeptide of 165 amino acids. An expression plasmid was constructed which permits the synthesis in E. coli of 2.5 x 10(8) units of interferon per litre of culture. This LeIF protected squirrel monkeys from lethal encephalomyocarditis virus infection.
511 citations
TL;DR: The alcohol dehydrogenase (ADH) gene (Adh) of Drosophila melanogaster was isolated by utilizing a mutant strain in which the Adh locus is deleted, and two intervening sequences were identified within Adh by S1 nuclease mapping experiments.
Abstract: The alcohol dehydrogenase (ADH; alcohol: NAD+ oxidoreductase, EC 1.1.1.1) gene (Adh) of Drosophila melanogaster was isolated by utilizing a mutant strain in which the Adh locus is deleted. Adult RNA from wild-type flies was enriched in ADH sequences by gel electrophoresis and then used to prepare labeled cDNA for screening a bacteriophage lambda library of genomic Drosophila DNA. Of the clones that hybridized in the initial screen, one clone was identified that hybridized with labeled cDNA prepared from a wild-type Drosophila strain but did not hybridize with cDNA prepared from an Adh deletion strain. This clone was shown to contain ADH structural gene sequences by three criteria: in situ hybridization, in vitro translation of mRNA selected by hybridization to the cloned DNA, and comparison of the ADH protein sequence with a nucleotide sequence derived from the cloned DNA. Comparison of the restriction site maps from clones of three different wild-type Drosophila strains revealed the presence of a 200-nucleotide sequence in one strain that was absent from the other two strains. The ADH mRNA sequences were located within the cloned DNA by hybridization mapping experiments. Two intervening sequences were identified within Adh by S1 nuclease mapping experiments.
475 citations
TL;DR: Double-stranded cDNA prepared from the 12S fraction of poly (A) RNA from interferon (IF)-producing human leukocytes was cloned in Escherichia coli using the pBR322 vector and one of the resulting clones had a 910-base pair insert which could hybridise to IF mRNA and was responsible for the production of a polypeptide with biological IF activity.
Abstract: Double-stranded cDNA prepared from the 12S fraction of poly(A) RNA from interferon (IF)-producing human leukocytes was cloned in Escherichia coli using the pBR322 vector. One of the resulting clones had a 910-base pair insert which could hybridise to IF mRNA and was responsible for the production of a polypeptide with biological IF activity. Up to 10,000 units IF activity per g of cells was obtained from some clones.
464 citations
TL;DR: The 12 Interferon (IFN)-related sequences detected in a human gene bank fall into not less than eight distinct classes, indicating that there are at least eight IFN-related genes.
Abstract: The 12 Interferon (IFN)-related sequences detected in a human gene bank fall into not less than eight distinct classes, indicating that there are at least eight IFN-related genes. Most, if not all, of these direct the synthesis of an IFN in Escherichia coli. The sequence of one chromosomal gene and its flanking regions was identical to that deduced for the cDNA corresponding to IFN-αl mRNA. No evidence was found for the existence of an intron, in either the coding or the non-coding segments of the gene.
432 citations
TL;DR: A cDNA library was constructed using mRNA from human fibroblasts induced with poly(I):poly(C) to create a bacterial clone containing fibroblast interferon cDNA sequences which is indistinguishable from authentic human Fibroblast Interferon by several criteria.
Abstract: A cDNA library was constructed using mRNA from human fibroblasts induced with poly(I):poly(C). A bacterial clone containing fibroblast interferon cDNA sequences was identified by hybridization to a cDNA probe synthesized using deoxyoligonucleotide primers which hybridize to fibroblast interferon mRNA specifically. Expression plasmids were constructed which permitted the synthesis in E. coli of 8 x 10(7) units of human fibroblast interferon per liter of culture. The bacterially produced fibroblast interferon is indistinguishable from authentic human fibroblast interferon by several criteria.
396 citations
TL;DR: The genes coding for the three membrane polypeptides of Semliki Forest virus have been sequenced and the primary structures of the proteins deduced and the amino acid sequence gives further insight into how the transmembrane structure of the three-chain virus membrane glycoprotein is generated in the infected cell.
Abstract: The genes coding for the three membrane polypeptides of Semliki Forest virus have been sequenced and the primary structures of the proteins deduced. The amino acid sequence gives further insight into how the transmembrane structure of the three-chain virus membrane glycoprotein is generated in the infected cell.
394 citations
TL;DR: A segment of a DNA encoding most of POMC is isolated, using as probe a mouse 144-base pair cloned cDNA fragment encoding β-MSH and β-endorphin, and this may indicate an as yet unrecognized biological function(s) for the NH2-terminal portion of the 16K fragment.
Abstract: The peptide hormones ACTH, beta-endorphin, alpha- and beta-melanotropin(MSH) and possibly gamma-MSH are synthesized in the pituitary gland by the processing of a 32,000-molecular weight (MW) polypeptide called proopiomelanocortin (POMC) The existence of a further precursor (pre form) to POMC containing an additional N-terminal 'leader' peptide has been suggested by analysis of the in vitro translation products of poly(A)-containing RNA from AtT-20 cells, a mouse ACTH-producing cell line of pituitary origin Nakanishi et al cloned and sequenced a cDNA copy of the bovine prePOMC mRNA This sequence confirmed the known structure of the carboxyl half of POMC and revealed the presence of a new MSH-like moiety, gamma-MSH, within the 16,000-MW amino half of the precursor (16K fragment) Recent experiments have suggested that this peptide may act in synergy with ACTH to increase corticosterone and aldosterone production in vivo and in vitro We have now isolated from a rat genomic DNA library a segment of a DNA encoding most of POMC, using as probe a mouse 144-base pair cloned cDNA fragment encoding beta-MSH and beta-endorphin The cloned rat gene is one of two (or more) closely related POMC genes The DNA sequence obtained shows that the cloned POMC gene is not interrupted by any intervening sequence (IVS) between the codon for amino acid 19 and the presumptive poly(A) addition site This region of POMC encodes all the biologically active peptides mentioned above The DNA sequence encoding the putative gamma-MSH and the coding sequence that precedes it are highly conserved between rat and cow This may indicate an as yet unrecognized biological function(s) for the NH2-terminal portion of the 16K fragment
296 citations
TL;DR: From the nucleotide sequence of the gene, the complete amino acid sequence of human fibroblast interferon was deduced and the protein is 166 amino acids long and is preceded by a 21-amino acid signal sequence.
Abstract: Chimaeric plasmids containing double-stranded cDNA copies of mRNA induced in human fibroblasts by poly I · C were screened by an RNA selection method. A series of clones to which human fibroblast interferon mRNA selectively hybridized was identified. From the nucleotide sequence of the gene, the complete amino acid sequence of human fibroblast interferon was deduced. The protein is 166 amino acids long and is preceded by a 21-amino acid signal sequence.
281 citations
TL;DR: Analysis of the βHCG cDNA nucleotide sequence suggests that this extension may have arisen by the loss of the termination codon of an ancestral β-like gene so that most of what was previously the 3′-untranslated region now codes for protein.
Abstract: A 579-base pair approximately full-length cDNA which codes for the 145-amino acid long beta-subunit of human chorionic gonadotropin (HCG) has been cloned in the plasmid vector pBR322 and its complete nucleotide sequence determined. A hydrophobic presequence of 20 amino acids can be identified from the nucleotide sequence. The amino acid sequence of the beta-subunit is known to be related to those of the beta-subunits of the other glycoprotein hormones LH, FSH and TSH, but the beta-subunit of HCG is unique in that it contains a C-terminal extension of about 30 amino acids which has no homologous counterpart in the other three hormones. Analysis of the beta HCG cDNA nucleotide sequence suggests that this extension may have arisen by the loss of the termination codon of an ancestral beta-like gene so that most of what was previously the 3'-untranslated region now codes for protein. The beta-subunit of HCG terminates with the codon UAA located 16 bases before the poly(A) in the sequence AAUAAA. This sequence is believed to be a recognition signal involved in either polyadenylation or processing and therefore has a dual role in this gene, serving both a coding and a regulatory function.
271 citations
TL;DR: The sequence of a human leukocyte-derived complementary DNA, Hif-2h, which directs the formation in Escherichia coli of a polypeptide, IFN-alpha 1, with interferon (IFN) activity has been described, revealing the presence of at least eightIFN-related genes.
Abstract: The sequence of a human leukocyte-derived complementary DNA (cDNA), Hif-2h, which directs the formation in Escherichia coli of a polypeptide, IFN-alpha 1, with interferon (IFN) activity has been described. A second IFN cDNA, Hif-SN206, which also elicits synthesis of a biologically active IFN, IFN-alpha 2, is described in this article. Whereas IFN-alpha 2 is twice as active on human as on bovine cells, IFN-alpha 1 is 10 to 20 times more active on bovine than on human cells. As deduced from the cDNA's, the messenger RNA's for the two IFN's differ in length and in 20 percent of the nucleotides; the mature IFN polypeptides differ in 17 percent of the amino acids. Both IFN-alpha 1 and IFN-alpha 2 differ from the lymphoblastoid IFN described by others. Therefore, at least three different IFN-alpha genes are expressed in man; studies on genomic DNA reveal the presence of at least eight IFN-related genes.
TL;DR: It was found that, in normal females, the genes for c36, c38, and at least one other chorion protein are specifically amplified more than 10-fold in the DNA of late-stage egg chambers, suggesting that altered gene dosage may be responsible for the decreased synthesis of chorions in the mutant.
Abstract: The endochorion and exochorion of Drosophila eggs are synthesized by the ovarian follicle cells during a brief period of about 5 hr. In this terminal phase of egg chamber development, the structural genes for several abundant chorion proteins are expressed at high levels according to a temporally regulated program. The female-sterile mutation ocelliless maps at the site of the genes for two of these proteins, the 36,000- and 38,000-dalton chorion proteins (c36 and c38), which are closely linked. The mutation results in a cis-acting reduction in the amounts of c36 and c38 that accumulate in late-stage egg chambers. We have investigated the mechanism that underlies this decreased production by using cDNA clones complementary to these gene sequences. Unexpectedly, it was found that, in normal females, the genes for c36, c38, and at least one other chorion protein are specifically amplified more than 10-fold in the DNA of late-stage egg chambers. The extra replication involves at least some adjacent chromosomal sequences and begins prior to the onset of mRNA and protein synthesis. The additional DNA remains stable after gene expression has ceased. The behavior of these genes is thus reminiscent of the properties of the DNA puffs that have been described in several groups of Diptera. The extent of amplification of c36 and c38, but not of the 18,000-dalton chorion protein c18 (which is unlinked), was decreased in the egg chambers of flies homozygous for ocelliless, suggesting that altered gene dosage may be responsible for the decreased synthesis of chorion proteins in the mutant.
TL;DR: Cell differentiation during A. nidulans asexual reproduction involves the accumulation of many new poly(A) RNA sequences, but not the loss of preexisting ones.
TL;DR: The data reported predict the hitherto unknown amino acid sequence of human fibroblast interferon and its putative signal peptide.
TL;DR: A strategy has been developed for sequencing the single-stranded RNA genes of influenza virus and the nucleotide sequence of segment 7 was completed and provides the primary structure of the matrix protein.
Abstract: A strategy has been developed for sequencing the single-stranded RNA genes of influenza virus. Restriction fragments derived from double-stranded cDNA copies of total influenza RNA were cloned into the bacteriophage M13mp2 and sequenced by the dideoxy technique. Sequences were extended and overlapped by using the virion RNA as template and priming with small restriction fragments. In the course of this strategy, the nucleotide sequence of segment 7 (1027 nucleotides) was completed and provides the primary structure of the matrix protein (27, 861 daltons). In addition, there is a second long reading frame which partly overlaps the reading frame of the matrix protein.
TL;DR: From clones cloned in the Escherichia coli plasmid pBR322, DNA encoding the mRNA for the precursor of the small subunit of the chloroplast enzyme ribulose-1,5-bisphosphate carboxylase is identified.
Abstract: Polyadenylated RNA from leaves of pea (Pisum sativum) has been copied into DNA and cloned in the Escherichia coli plasmid pBR322. From these clones we have identified and sequenced DNA encoding the mRNA for the precursor of the small subunit of the chloroplast enzyme ribulose-1,5-bisphosphate carboxylase.
TL;DR: Cloned DNA complementary to the messenger RNAs for four immunologically distinct VSGs are cloned and hybridised these complementary DNAs with restriction digests of T. brucei nuclear DNA and infer that activation of a VSG gene involves the production of an expression-linked copy of that gene.
Abstract: Pathogenic African trypanosomes evade the immune system of their mammalian hosts by the sequential expression of alternative cell-surface glycoproteins (reviewed in refs 1,2). Variant surface glycoproteins (VSGs) purified from cloned variants of Trypanosoma brucei have similar molecular weights (about 60,000), but differ in amino acid composition, N-terminal amino acid sequence and C-terminal structure. We have cloned DNA complementary to the messenger RNA's for four immunologically distinct VSGs and hybridised these complementary DNAs (cDNAs) with restriction digests of T. brucei nuclear DNA, fractionated by gel electrophoresis and transferred to nitrocellulose strips. Each cDNA recognises a unique set of fragments and this basic set is present unaltered in the nuclear DNAs from the four variants. In addition, each probe recognises an extra fragment only in nuclear DNA isolated from cells expressing the VSG corresponding to the cDNA probe. We infer that activation of a VSG gene involves the production of an expression-linked copy of that gene.
TL;DR: The results and quantitative aspects of the selection procedure suggest that developmentally controlled poly(A)+ RNA coding regions may not be distributed randomly in the Aspergillus genome.
TL;DR: Double-stranded cDNA was synthesized from a mouse liver mRNA fraction enriched for metallothionein mRNA activity, ligated to restriction site linkers, inserted into pBR322, and used to transform Escherichia coli chi 1776 to screen both a mouse myeloma and a mouse embryo DNA library in bacteriophage lambda.
Abstract: Double-stranded cDNA was synthesized from a mouse liver mRNA fraction enriched for metallothionein mRNA activity, ligated to restriction site linkers, inserted into pBR322, and used to transform Escherichia coli chi 1776. The sequence of the largest plasmid containing DNA that hybridized to metallothionein mRNA was determined and shown to contain a 380-base-pair insert that includes the entire coding region and 3' untranslated region of metallothionein-I. The metallothionein-I insert was nick-translated and used to screen both a mouse myeloma and a mouse embryo DNA library in bacteriophage lambda. A metallothionein-I genomic clone containing 13-15 kilobase pairs of mouse DNA was isolated from each library. Both contain a 3.8-kilobase-pair EcoRI fragment that hybridizes to the metallothionein-I probe. The location, size, and orientation of the metallothionein-I gene within the 3.8-kilobase-pair fragment were determined by heteroduplex and restriction mapping. The gene spans 1.1 kilobase pairs and contains at least two introns.
TL;DR: Several genes for the variable region of immunoglobulin heavy chains (VH genes) have been isolated from human fetal liver DNA by using a cDNA plasmid probe containing a mouse VH sequence, suggesting that similar enzymatic mechanisms are involved in the integration of V genes in both heavy and light chains.
Abstract: Several genes for the variable region of immunoglobulin heavy chains (VH genes) have been isolated from human fetal liver DNA by using a cDNA plasmid probe containing a mouse VH sequence. The detectable VH genes are separated by 12-16 kilobases of DNA, and hybridization experiments show about 23 hybridizing VH genes in DNA of three different individuals. The complete nucleotide sequence of one of these human VH genes shows that it belongs to the human VHIII subgroup. The VH gene appears to contain an intervening sequence (104 bases in length) within a precursor sequence, between residues -4 and -5. The precursor sequence is itself 19 codons in length. The 3' end of the V gene seems to be at codon 93 or 94, and this is followed by the conserved sequences C-A-C-A-G-T-G and G-A-C-A-C-A-A-A-C-C. The presence of these sequences suggests that similar enzymatic mechanisms are involved in the integration of V genes in both heavy and light chains.
TL;DR: Nucleotide sequence analysis determined the sequence of the 26-amino acid signal peptide and confirmed the published amino acid sequence ofThe secreted hormone at all but 2 residues, and the nucleotide sequence of bGH mRNA is 83.9% homologue with rat GH mRNA and 76.5% homologous with human GH mRNA.
TL;DR: The results indicate that salivary gland and liver amylase mRNAs are transcribed from identical or very closely related genes which differ from that expressed in the pancreas.
TL;DR: Preliminary Southern blot analysis of Eco RI restriction fragments of mouse embryo DNA revealed very complex patterns, suggesting that a family of homologous sequences for the individual rp-mRNAs may exist in the mouse genome.
TL;DR: The apo VLDLII and vitellogenin genes appear to be the only genes capable of high rates of expression in the liver that exhibit such an exceptional response to the hormone.
Abstract: Livers of egg-laying species contain abundant mRNAs encoded by both estrogen-responsive and constitutively expressed genes. We have recently constructed cDNA clones from three members of the abundant mRNA class of hen liver. One of these mRNA species was identified as serum albumin mRNA, and another as vitellogenin mRNA. In this study we have identified the third member of the group as apo VLDLII mRNA. Hybridization analyses using cloned cDNA probes indicate that expression of the apo VLDLII gene in rooster liver, like that of the vitellogenin gene, is completely dependent upon the administration of estrogen. The apo VLDLII and vitellogenin genes appear to be the only genes capable of high rates of expression in the liver that exhibit such an exceptional response to the hormone. Administration of estrogen resulted in the appearance of both mRNA species within 30 min, followed by a rapid accumulation to several thousand copies per cell. Removal of the hormone caused a marked destabilization of both vitellogenin mRNA and apo VLDLII mRNA. In contrast, the absolute levels of serum albumin mRNA were unaffected by the hormone. Comparative studies on the structure and organization of these three genes may reveal elements involved in determining their rates of expression in the presence and absence of estrogen.
TL;DR: The genomic organization of the mouse dihydrofolate reductase gene has been determined by hybridization of specific cDNA sequences to restriction endonuclease-generated fragments of DNA from methotrexate-resistant S-180 cells, indicating that this organization represents that of the natural gene.
TL;DR: A human genomic DNA segment encoding the corticotropin-beta-lipotropin precursor peptide from a fetal DNA library is isolated, using previously cloned bovine cDNA for this peptide as a probe.
Abstract: We have isolated a human genomic DNA segment encoding the corticotropin-beta-lipotropin precursor peptide from a fetal DNA library, using previously cloned bovine cDNA for this peptide as a probe. The human genomic DNA was studied by electron microscope heteroduplex analysis and gel blotting methods, and its nucleotide sequence was determined and compared with that of cDNA corresponding to bovine pro-opiomelanocortin mRNA. From this sequence, segments of interspecies conservation and divergence, punctuated by pairs of the basic amino acid residues lysine and arginine, were identified. No noncoding intervening sequence was observed over an 830-base-pair DNA segment extending from a position near the 5' end of the structural pro-opiomelanocortin gene through the 3' terminus of the cDNA and including sequences for the component peptide hormones corticotropin and beta-lipotropin.
TL;DR: The nucleotide sequence of the human leukocyte interferon cDNA carried in hybrid plasmid Z-pBR322(Pst)/HcIF-2h, which has been shown to direct the formation of a polypeptide with human leucocyteinterferon activity, suggests that these two interferons are encoded by two non-allelic genes.
TL;DR: In bacteria, recombinant plasmids containing DNA complementary to mRNA coding for a pancreatic pre-prosomatostatin, a product of the cell-free translation of pancreatic islet mRNAs, are constructed and cloned, suggesting that this particular sequence may have biologic importance in cellular processing of the som atostatin precursor.
Abstract: We have constructed and cloned, in bacteria, recombinant plasmids containing DNA complementary to mRNA coding for a pancreatic pre-prosomatostatin, a product of the cell-free translation of pancreatic islet mRNAs shown previously by immunoprecipitation to be a precursor of somatostatin. A clone containing a nearly full-length cDNA insert of 550 base pairs was identified and appeared to contain the entire coding sequence for the somatostatin precursor in addition to portions of the 5' and 3' untranslated regions. mRNA coding for the pre-prosomatostatin is 600-630 bases long as determined by agarose gel electrophoresis and hybridization with labeled cDNA. Analyses of the nucleotide sequence of the cDNA revealed a protein of 119 amino acid beginning with methionine followed by a typical leader sequence containing 18 hydrophobic amino acids. The tetradecapeptide somatostatin, identical in sequence to mammalian hypothalamic somatostatin, is located at the carboxy terminus followed immediately by a stop codon. An ARg-Lys sequence immediately preceding the sequence of somatostatin is typical of a prohormone cleavage site. A sequence Ala-Pro-Arg-Glu preceding the Arg-Lys cleavage site is identical to that found in porcine prosomatostatin. The evolutionary conservation of the identical amino acid sequence of the somatostatin tetradecapeptide from fish to mammals is remarkable. In addition, similar conservation, in fish and mammals, of the sequence Ala-Pro-Arg-Glu-Arg-Lys preceding the coding region for somatostatin suggets that this particular sequence may have biologic importance in cellular processing of the somatostatin precursor.
TL;DR: Restriction of phi X174 DNA have led to the identification of a 55-nucleotide fragment that carries the protein n' recognition sequence, a sequence of 44 nucleotides that forms a stable hairpin structure that may be the signal for protein n', a prepriming DNA replication enzyme of Escherichia coli, to initiate the pre priming events that led toThe start of pho X174 complementary DNA strand replication.
Abstract: Protein n', a prepriming DNA replication enzyme of Escherichia coli, is a phi X174 DNA-dependent ATPase. Restriction of phi X174 DNA have led to the identification of a 55-nucleotide fragment that carries the protein n' recognition sequence. Molecular hybridization and sequence analysis have located this sequence within the untranslated region between genes F and G, a map location analogous to that of the unique complementary strand origin of phage G4 DNA. Within the 55-nucleotide fragment is a sequence of 44 nucleotides that forms a stable hairpin structure. This duplex may be the signal for protein n' to initiate the prepriming events that led to the start of phi X174 complementary DNA strand replication.
TL;DR: Analysis of two different plasmids containing pituitary cDNA inserts has provided the complete 681-nucleotide coding sequence of preprolactin, suggesting that the intervening sequences of the rat prolactin gene contain DNA sequences which are repeated elsewhere in the rat genome.