Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “DNA, Complementary”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

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

At least 37 records · Page 2Linked to original sources
PubMed1977

Measurement of proviral genes in uninfected and avian myeloblastosis virus-infected cells by hybridization with 3H-labeled complementary DNA probe excess.

Viral RNA (vRNA) from avian myeloblastosis virus or DNA from virus-infected and uninfected cells was hybridized with [3H]DNA complementary to viral RNA ([3H]cDNA) under conditions of [3H]cDNA excess. When [3H]cDNA was used to drive the hybridization reaction with vRNA, a rate constant of 33.2 liters/mol-s was obtained. The same rate constant was obtained when vRNA excess was used as the driver. The specific activities of the [3H]DNA probe, estimated from kinetic measurements of the hybridization reaction and from the amount of [3H]cDNA in hybrid form at equilibrium, were 9.1 and 8.6 cpm/pg, respectively. DNA isolated from uninfected cells contained five or six copies of proviral DNA per cell genome. DNA isolated from erythrocytes infected with avian myeloblastosis virus had an additional five or six viral genes added to the cell genome, and the virus-infected target cell (myeloblasts) contained about 15 additional copies of proviral DNA per cell. The use of excess [3H]cDNA probe is an easy and accurate method to quantify the frequency of proviral DNA sequences in cell DNA and to measure a small amount (40 to 200 pg) of vRNA. Probe excess hybridization offers a number of advantages over other procedures and these are discussed.

Animals↗
PubMed1980

Nucleotide sequence of human preproinsulin complementary DNA.

Recombinant bacterial plasmids that contain DNA complementary to human preproinsulin messenger RNA have been constructed. One clone contains the entire preproinsulin coding region, as well as the 3' untranslated region of the messenger RNA and eight nucleotides of the 5' untranslated region. Additional sequence information for the 5' untranslated region was obtained with the use of insulinoma messenger RNA in conjunction with specific primers from the cloned DNA for enzymatic chain termination sequence analysis. The results confirm the amino acid sequence of human proinsulin previously determined, and predict the amino acid sequence of the human preproinsulin signal peptide.

Amino Acid Sequence↗
PubMed1976

Effects of prolactin and progesterone on expression of casein genes. Titration of casein mRNA by hybridization with complementary DNA.

Prolactin injected into pseudopregnant rabbits induced lactogenesis. Progesterone injected simultaneously considerably reduced this effect. The content of mRNA for casein in total cellular RNA and in total polysomal RNA was estimated during these two hormonal actions using a hybridization technique. The synthesis of DNA complementary to casein mRNA was carried out with mRNA obtained from polysomes immunoprecipitated with anti-casein antibody. The content of casein mRNA was determined by hybridization of this cDNA with the RNA fractions. This technique revealed that prolactin induced the synthesis of mRNA for casein whereas progresterone greatly limited this effect of prolactin. These variations of casein mRNA content are in good agreement with values previously determined by translation and with the ability of mammary gland to synthesize casein.

Animals↗
PubMed1981

The preparation and characterization of locust vitellogenin messenger RNA and the synthesis of its complementary DNA.

Poly(A)+ (polyadenylated) RNA was isolated from vitellogenic female-locus fat-body by LiCl/urea extraction and poly(U)-Sepharose 4B affinity chromatography. Agarose-gel electrophoresis of this poly(A)+ RNA under denaturing conditions shows the presence of a high-molecular-weight species (greater than 31 S, 7100 nucleotides) as the major species, which is absent from the RNA prepared from male-locust fat-body. Inclusion of this poly(A)+ RNA in a mRNA-dependent reticulocyte-lysate system directs the synthesis of polypeptides that could be immunoprecipitated with monospecific antibodies against locust egg vitellin. DNA complementary (cDNA) to the poly(A)+ RNA was synthesized, and back-hybridization of the cDNA to its template reveals a major abundant species comprising about 45% of the total poly(A)+ RNA hybridizing with R0t 1/2 of 2 x 10(-2) mol . litre-1 . s. Abundant cDNA isolated from the total cDNA hybridizes to poly(A)+ RNA with a R0t 1/2 of 9 x 10(-3) mol . litre-1 . s. There are 9.1 x 10(3) copies of vitellogenin mRNA per cell of vitellogenic female-locust fat-body, comprising 55% of the poly(A)+ RNA and equivalent to 0.7% of total cellular RNA.

Animals↗
PubMed1979

Human growth hormone: complementary DNA cloning and expression in bacteria.

The nucleotide sequence of a DNA complementary to human growth hormone messenger RNA was cloned; it contains 29 nucleotides in its 5' untranslated region, the 651 nucleotides coding for the prehormone, and the entire 3' untranslated region (108 nucleotides). The data reported predict the previously unknown sequence of the signal peptide of human growth hormone and, by comparison with the previously determined sequences of rat growth hormone and human chorionic somatomammotropin, strengthens the hypothesis that these genes evolved by gene duplication from a common ancestral sequence. The human growth hormone gene sequences have been linked in phase to a fragment of the trp D gene of Escherichia coli in a plasmid vehicle, and a fusion protein is synthesized at high level (approximately 3 percent of bacterial protein) under the control of the regulatory region of the trp operon. This fusion protein (70 percent of whose amino acids are coded for by the human growth hormone gene) reacts specifically with antibodies to human growth hormone and is stable in E. coli.

Amino Acid Sequence↗
PubMed1983

Synthesis and characterization of a DNA complementary to Xenopus laevis albumin mRNA.

Albumin complementary DNA (cDNA) was transcribed from purified albumin mRNA from the liver of Xenopus laevis. The resultant cDNA was an almost full length copy as defined by denaturing gel electrophoresis; was hybridized specifically to albumin mRNA with pseudo-first-order reaction kinetics and the mRNA . cDNA hybrid exhibited a sharp melting profile with a Tm of 83 degrees C. The identity of the cDNA was confirmed by gel electrophoresis following hybridization-arrested translation.

Albumins↗
PubMed1979

Synthesis and characterization of a DNA complementary to pre-uteroglobin mRNA.

Uteroglobin, a progesterone-induced uterine protein of the rabbit, is synthesized in cell-free systems as a precursor containing 21 additional amino-acids at its N-terminal end. The mRNA for pre-uteroglobin has been purified from the membrane-bound polysomes of induced endometrium and used as template for the synthesis of a full copy complementary DNA. Final purification of the cDNA was based on hybridization to the template mRNA up to a low value of r0t (0.01 M . s) and digestion of the non-hybridized cDNA by S1 nuclease. A comparison of the hybridization kinetics of the pre-uteroglobin cDNA and rabbit globin cDNA to their respective templates indicates a nucleotide sequence complexity of 650 for pre-uteroglobin mRNA, in agreement with the values obtained by sucrose gradient centrifugation and polyacrylamide gel electrophoresis in formamaide. The melting temperature of the hybrids of pre-uteroglobin cDNA to its template reflects the absence of mismatched sequences. This cDNA has been used to quantify pre-uteroglobin mRNA sequences in the endometrial RNA from control animals and from animals treated sequentially with estradiol and progesterone. In agreement with the induction of uteroglobin-synthesizing activity, there is a dramatic increase in the uterine content of pre-uteroglobin mRNA after hormonal treatment. Part of this effect can be accounted for by hormonally induced cell proliferation. When expressed on a DNA basis there is a 50--100-fold increase in the cellular content of pre-uteroglobin mRNA following hormonal treatment.

Animals↗
PubMed1992

Isolation and sequence of sheep immunoglobulin E heavy-chain complementary DNA.

A complementary DNA (cDNA) corresponding to sheep immunoglobulin E heavy (IgE H) chain messenger RNA (mRNA) was isolated and sequenced. A fragment of sheep IgE constant H (epsilon)-chain was initially synthesized using the polymerase chain reaction (PCR) and single-stranded cDNA prepared from the mRNA of parasite-stimulated sheep lymph nodes. This fragment was then used to probe a cDNA library, again prepared from a parasite-stimulated lymph node. A recombinant clone containing cDNA encoding a sheep IgE H-chain of 1802 base pairs was isolated and the H-chain cDNA-sequenced. The nucleotide sequence was found to code for a complete sheep IgE H-chain, consisting of both variable (V) and constant (C) regions. When the amino acid sequence derived from the nucleotide sequence was compared to other species epsilon-chains, interesting homologies and differences between corresponding domains were found, including conservation in cysteine and tryptophan residues and variable glycosylation sites.

Amino Acid Sequence↗
PubMed1977

Human placental lactogen mRNA and its structural genes during pregnancy: quantitation with a complementary DNA.

A complementary DNA (cDNA) strand was transcribed from human placental lactogen (hPL) mRNA. Based on alkaline sucrose gradient centrifugation, the size of the cDNA was about 8 S, which would represent at least 80% of the hPL mRNA. Previously we showed that four to five times more hPL was synthesized in cell-free extracts derived from term as compared to first trimester placentas. Hybridization of the cDNA with RNA derived from placental tissue revealed that there was about four times more hPL mRNA sequences in total RNA from term placenta than in a comparable quantity of total first trimester RNA. Only background hybridization was observed when the cDNA was incubated with RNA prepared from human kidney. To test if this differential accumulation of hPL mRNA was the result of an amplification of hPL genes, we hybridized the labeled cDNA with cellular DNA from first trimester and term placentas and with DNA isolated from human brain. In all cases, the amount of hPL sequences was approximately two copies per haploid genome. Thus, the enhanced synthesis of hPL mRNA appears to result from a transcriptional activation rather than an amplification of the hPL gene. The increase likely reflects placental differentiation in which the proportion of syncytial trophoblast increases at term.

DNA↗
PubMed1987

Complete amino acid sequence of human ornithine decarboxylase deduced from complementary DNA.

A complementary DNA (cDNA) encoding ornithine decarboxylase was isolated from a human liver cDNA library, and the nucleotide sequence coding for the entire enzyme was determined. The 1825-nucleotide-long cDNA contained an open reading frame of 1383 nucleotides, 87 nucleotides 5' from the first methionine codon, 346 nucleotides in the 3'-noncoding region, and a poly(A) tail of nine bases. Primer extension studies indicated that the 5'-noncoding region of the human ornithine decarboxylase mRNA was 335 nucleotides long. The amino acid sequence deduced from the open reading frame for a 461-residue polypeptide predicts a molecular weight of 51.156 for the human enzyme and has about 90% homology with the amino acid sequence of the murine ornithine decarboxylase (44 differences among the 461 amino acids). The nucleotide sequences of the human and murine ornithine decarboxylase mRNAs share an 85% homology, even in their 3'-noncoding regions. In contrast to rodent tissues with two ornithine decarboxylase mRNAs, normal human tissues appear to express only a single mRNA species with a molecular size of 2.25 kb. Southern blotting of human leukocyte DNA from 20 individuals indicated that the ornithine decarboxylase gene belongs to a multigene family in man and showed restriction fragment length polymorphism when cleaved with Pst I, but not when cleaved with Pvu II, Msp I, Hinc II, or Bam HI.

Amino Acid Sequence↗
PubMed1989

Identification of the cystic fibrosis gene: cloning and characterization of complementary DNA.

Overlapping complementary DNA clones were isolated from epithelial cell libraries with a genomic DNA segment containing a portion of the putative cystic fibrosis (CF) locus, which is on chromosome 7. Transcripts, approximately 6500 nucleotides in size, were detectable in the tissues affected in patients with CF. The predicted protein consists of two similar motifs, each with (i) a domain having properties consistent with membrane association and (ii) a domain believed to be involved in ATP (adenosine triphosphate) binding. A deletion of three base pairs that results in the omission of a phenylalanine residue at the center of the first predicted nucleotide-binding domain was detected in CF patients.

Amino Acid Sequence↗
PubMed1975

Induction of ovalbumin mRNA sequences by estrogen and progesterone in chick oviduct as measured by hybridization to complementary DNA.

A complementary DNA synthesized from ovalbumin mRNA was used in hybridization experiments to study the early effect of estrogen and progesterone on the accumulation of ovalbumin mRNA sequences in the chick oviduct. Chicks treated with estrogen withdrawn from the hormone maintain a steady level of 60 molecules of ovalbumin mRNA per tubular gland cell, at least 80% of which are localized in the cytoplasm. After estrogen administration, there is a 3- to 4-hour lag before a rapid increase in the number of ovalbumin mRNA sequences and a parallel increase in ovalbumin synthesis. Progesterone causes a more rapid increase in both ovalbumin mRNA sequences and ovalbumin synthesis with a lag period of only 90 min. The hybridization results demonstrate that both estrogen and pregesterone affect the amount of ovalbumin mRNA per cell. The 3-hour lag period seen with estrogen appears to be caused by some event after the binding of the estrogen receptor to chromatin but prior to change in the rate of transcription of the ovalbumin gene.

Animals↗
PubMed1985

Characterization of genomic and complementary DNA sequence of human C-reactive protein, and comparison with the complementary DNA sequence of serum amyloid P component.

Complementary and genomic DNA clones corresponding to the human C-reactive protein (CRP) mRNA and structural gene have been analyzed and compared. Nucleotide sequencing of the coding regions of both cDNA and genomic DNA revealed an additional 19 amino acid peptide not described in the published CRP amino acid sequence. The CRP gene contains a single 278 base pair intron within the codon specifying the third residue of mature CRP. The intron contains a repetitive sequence (GT)15G(GT)3 which is similar to structures capable of adopting the Z-DNA form. A comparison of CRP coding and amino acid sequences with those of serum amyloid P component revealed striking overall homology which was not uniform: a region of limited conservation is bounded by two highly conserved regions.

Amino Acid Sequence↗
PubMed1998

The detection of Trichinella with polymerase chain reaction (PCR) primers constructed using sequences of random amplified polymorphic DNA (RAPD) or sequences of complementary DNA encoding excretory-secretory (E-S) glycoproteins.

Diagnostic PCR primers for Trichinella were constructed. Twelve pairs of primers were designed based on the sequences of random amplified polymorphic DNA, and 4 pairs of primers were designed based on the reported sequences of complementary DNA encoding excretory-secretory glycoproteins. With these primers, 31 samples of DNA from different strains of Trichinella including 5 species (Trichinella spiralis, Trichinella britovi, Trichinella nativa, Trichinella nelsoni and Trichinella pseudospiralis) and 3 phenotypes of uncertain taxonomic level (Trichinella T5, T6 and T8) were tested with PCR. Genus Trichinella can be identified by 4 different primer pairs (SB147D, SB372A, SB153, or Ts43). Trichinella spiralis can be identified by the presence of a 673 bp amplicon in PCR with the primer pair SB147B. Trichinella nelsoni can be identified using primer pair SB147F or by the presence of 673 bp and ca. 380 bp amplicon in PCR with the primer pair SB147B. Trichinella pseudospiralis can be identified by 2 primer pairs (SB147E or SB372B). Trichinella T5 can be identified by the primer pair SB147G. Trichinella T8 can be identified by its positivity by the primer pair SB147C and its negativity by the primer pair SB372C. A group of Trichinella species (T. britovi, T. nativa and Trichinella T6) can be identified by the primer pair SB372C.

Animals↗
PubMed1995

Large-scale complementary DNA sequencing methods.

Complementary DNA libraries are useful tools for uncovering genes of interest in C. elegans and finding specific homologies to genes in other organisms (Waterston et al., 1992; McCombie et al., 1992). When working with existing cDNA libraries, be sure to carefully choose which libraries would be most beneficial to the type of research being done. Some libraries may be specific for genes that are present in lower copy numbers, whereas others may be of a more general nature. It is important to fully understand the source and construction of the library you will be working with. Once an appropriate library has been chosen, work may begin to isolate a specific cDNA and sequence it completely or to survey many cDNAs by single-pass DNA sequencing. Whatever the project, it is important to develop a specific strategy for both the sequencing and the organization of the clones being characterized. The strategies and procedures we have outlined in this chapter have proven effective for rapid and comprehensive cDNA characterization.

Animals↗
PubMed2004

Molecular cloning and characterization of a maize transglutaminase complementary DNA.

Two related complementary DNA clones, TGZ15 and TGZ21, encoding active maize transglutaminase (TGase) have been isolated for the first time in plants by molecular cloning (Patent Pending PCT/ES03/00247). Southern and northern blot analyses indicate that the two cDNAs probably corresponded to two different single-copy genes in the maize genome. Northern blot analyses revealed that the transcript is expressed preferentially in young leaves and differentiated embryogenic maize callus. This expression is dependent on light exposure time. TGase activity of the proteins encoded by clones TGZ15 and TGZ21 was detected in bacterial extracts overexpressing them, using two enzymatic assays. TGase activity was significantly higher than that of the empty-phagemid bacterial extracts. As in other TGases, this activity was inhibited by monodansyl cadaverine (MDC), GTP and the absence of exogenous Ca(2+). Likewise, light-stimulated Ca(2+)-dependent TGase activity was detected in thylakoids and grana of maize chloroplast, which was inhibited by MDC, GTP, DIECA and Diuron.

Amino Acid Sequence↗
PubMed1976

Fractionation of DNA nucleotide transcripts from Moloney sarcoma virus and isolation of sarcoma virus-specific complementary DNA.

Radioactive DNA complementary to nucleotide sequences in Moloney murine sarcoma virus (MSV) and Moloney leukemia virus (M-MuLV) complex was made by the endogenous reverse transcriptase reaction. These virus stocks contained a threefold excess of MSV over M-MuLV as measured by biological assay. The complementary DNA was an accurate copy of the viral RNA in that 86% of 35S viral RNA hybridized with complementary (cDNA) DNA at a 1.5 to 1 cDNA-RNA molar ratio. The complementary DNA, of a 4-6S size, was fractionated by sequential absorptions with MulV and the feline leukemia virus pseudotype of MSV, [MSV(FeLV)] RNA. In this manner three sets of nucleotide sequences whichrepresent different portions of the MSV viral complex were obtained: a sarcoma virus-specific fraction (cDNAsarc) with sequences that had no homology to M-MuLV RNA but which hybridized to MSV (FeLV) RNA, a sarcoma-leukemia fraction (cDNA common) with sequences common to MSV as well as M-MuLV viral RNA, and a cDNAleuk representing those nucleotide sequences found only in M-MuLV. Hybridization of MSV-MuLV viral 35S RNA with a threefold molar excess of cDNA's revealed that approximately 20% was hybridized with cDNAsarc, whereas approximately 75% was hybridized with cDNAcommon. M-MuLV 35S RNA alone did not hybridize with cDNAsarc but did hybridize 40 and 50% with cDNAleuk and cDNAcommon, respectively. The cDNAsarc represents about 25% of the total MSV sequences, whereas the cDNAcommon represents the remainder of the MSV virus genome. Some cDNAcommon sequences were shared by two other sarcoma viruses and several distinctly different isolates of MulV. In contrast, the MSV "sarc" sequences had little or no homology with two other murine sarcoma virus isolates.

Base Sequence↗
PubMed1988

Complementary DNA cloning of cytochrome P-450s related to P-450(M-1) from the complementary DNA library of female rat livers. Predicted primary structures for P-450f, PB-1, and PB-1-related protein with a bizarre replacement block and their mode of transcriptional expression.

Three kinds of cDNA clones were isolated from a cDNA library synthesized from female rat liver mRNA by cross-hybridization with the P-450(M-1) cDNA as a probe and sequenced. One clone appears to be the previously isolated P-450f cDNA clone with an additional 5'-untranslated and coding sequence which are lacking in the previously reported clone (Gonzalez, F. J., Kimura, S., Song, B.-J., Pastewka, J., Gelboin, H. V., and Hardwick, J. P. (1986) J. Biol. Chem. 261, 10667-10672), though several nucleotide differences were seen. Another one is for P-450PB-1 mRNA previously isolated, and the last has an almost identical nucleotide sequence to P-450PB-1 (the same report cited above) except for one region of 159 base pairs where the sequence homology between the two is abruptly broken down. This nonhomologous region appears to correspond exactly to the entire eighth exon, estimated by comparison with the gene structure of the related P-450 (P-450(M-1)). This replacement in P-450PB-1 (ps) causes a frameshift in the open reading frame, resulting in the generation of a truncated form of P-450 with a strange replacement block and lacking the heme-binding region. This observation suggests that the mRNA whose cDNA was cloned here was produced from a recombinant gene generated by gene conversion or from alternative splicing of a cryptic exon. Sex- and age-dependent expression of the mRNAs investigated by dot blot analysis revealed that normal- and pseudo-type PB-1 mRNA were expressed in both male and female rat livers, though their age-dependent expression was different in male and female animals. In addition, both the mRNAs were specifically expressed in the female brain of 8 weeks, whereas practically no expression was observed in kidney and lung of both sexes.

Amino Acid Sequence↗