Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Complementary DNA (cDNA) was prepared with RNA-dependent DNA polymerase from human globin messenger RNA (mRNA). Annealing and translation experimenta with total mRNA from circulating cells from a patient with heterozygous beta/heterozygous beta-delta-o thalassemia (beta-o/delta beta-o-thalassemia) demonstrated no detectable mRNA for beta-globin. cDNA enriched in sequences homologous to beta-globin mRNA was prepared by hydroxylapatite fractionation of hybrids formed between beta-o/delta beta-o-thalassemic mRNA and cDNA made from mRNA from a patient with alpha-thalassemia (hemoglobin H disease). The rate of annealing of this beta-enriched cDNA to normal human nuclear DNA was that of a sequence present as only a single copy per haploid genome. The beta-enriched cDNA annealed to the beta-o-delta beta-o-thalassemia total DNA with approximately the same kinetics as to normal DNA, indicating that no total gene deletion of beta-globin genes from the diploid genome has occurred, although the accuracy of the technique could not exclude with certainty a partial deletion or a deletion of a beta-globin gene from only one of the haploid genomes. This demonstrates that at least one of the beta-o- or the delta beta-o-thalassemia haploid genomes in this case contains a substantially intact beta-globin gene.
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PMID:Human globin gene analysis for a patient with beta-o/delta beta-thalassemia. 4 57

The 5' noncoding region sequence of rabbit beta-globin mRNA has been determined. This region is 53 nucleotides long, not including the A-U-G initiator sequence or m7Gppp "cap" structure. 32P-labeled DNA complementary to the 5' noncoding region was synthesized using reverse transcriptase with the synthetic deoxyoctanucleotide d(G-C-A-C-C-A-T-T) as a "primer". The cDNA produced was then sequenced using a modification of the gel-sequencing technique previously developed for DNA sequencing (G.G. Brownlee and E.M. Cartwright, manuscript in preparation). The sequence obtained was checked by depurination and nearest-neighbor analysis. The known sequence at the 3' end of rabbit 18S ribosomal RNA cannot base-pair extensively with the 5' noncoding region of beta-globin mRNA; however, it does form 6 base pairs around the initiation codon.
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PMID:Complete nucleotide sequence of the 5' noncoding region of rabbit beta-globin mRNA. 6 96

Rabbit globin complementary DNA made with RNA-dependent DNA polymerase (reverse transcriptase) was used as a template for in vitro synthesis of 32P-labeled RNA and deoxysubstituted RNA. The sequences of the nucleotides in most of the fragments resulting from combined ribonuclease T1 and alkaline phosphatase digestion have been determined. In addition, the 3' nearest neighbor was determined for several fragments resulting from digestion with T1 ribonuclease. The utility of the deoxysubstitution technique was demonstrated by the ease with which the sequences of pyrimidine-rich fragments could be determined. Many sequences thus determined were long enough to fit uniquely with the alpha- or beta-globin amino acid sequences. The positions of these fits were found to be clustered, leading us to believe that only certain regions of the complementary DNA are transcribed by Escherichia coli RNA polymerase. Other unique characteristics of RNA synthesis from a complementary DNA template include a high yield of free poly(A) and the fact that one must use low rather than high salt buffers to obtain transcripts of high molecular weight.
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PMID:Rabbit globin mRNA: analysis of T1 RNAse digestion fragments. 6 35

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.
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PMID:Human beta-globin messenger RNA. III. Nucleotide sequences derived from complementary DNA. 6 58

Rabbit and human alpha-globin complementary DNA was synthesized using the primer (dT10)dGdCdC hybridized to globin mRNA with reverse transcriptase, and sequenced using the plus and minus gel sequencing procedure (Sanger and Coulson, 1975; Brownlee and Cartwright, 1977). The complete 3' noncoding region, 89 nucleotides in length, and one third of the coding region of rabbit alpha-globin mRNA have thus been sequenced. These data are compared with the near complete 3' noncoding region sequence of human alpha-globin mRNA obtained in these and previous studies. The two mRNAs are approximately 80% homologous in their 3' noncoding regions, except that the human sequence has an insert of 24 nucleotides. A very similar insert in sequence has been described in human beta-globin mRNA (Proudfoot, 1977).
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PMID:Nucleotide sequence of the 3' terminal third of rabbit alpha-globin messenger RNA: comparison with human alpha-globin messenger RNA. 7 Feb 77

We have found that double-stranded cDNA synthesized in extended reactions by avian myeloblastosis virus reverse transcriptase is suitable substrate for a variety of restriction endonucleases. Experiments in which rabbit reticulocyte mRNA was reverse-transcribed and restricted to generate beta-globin-specifihe nucleotide sequence of beta-globin mRNA. This method has been applied to study collagen mRNA synthesis in normal and Rous sarcoma virus (RSV)-transformed chick embryo fibroblasts. Characteristic sets of collagen cDNA restriction fragments were produced from RNA fractions rich in collagen message activity. These sets of cDNA fragments, generated by the restriction endonucleases Hae III and Hap II, provided a convenient marker for the presence of collagen mRNA sequences. Equal quantities of high molecular weight mRNA from chick embryo fibroblasts (CEF) and RSV-CEF were reverse-transcribed and the resulting cDNA was restricted. The relative yields of collagen cDNA fragments from such reactions strongly suggest that the decrease in functional collagen RNA following RSV-induced transformation of CEF represents a decrease in the copy number of collagen messenger sequences. The potential of this approach for the study of regulation in other systems is discussed.
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PMID:Decreased levels of collagen mRNA in rous sarcoma virus-transformed chick embryo fibroblasts. 7 27

The nucleotide sequence of the entire 5' untranslated region of human gamma-globin mRNA has been determined. This was accomplished by analyzing complementary DNA (cRNA) synthesized from the mRNA with reverse transcriptase. The CDNA was labeled at its 3' end with 32"p using terminal deoxynucleotidyl transferase, digested with the restriction endonuclease Hae III and the end-labeled fragment isolated ans sequenced by the method of Maxam and Gilbert. Including the initiation codon AUG, the 5' untranslated region of human gamma-globin mRNA contains 57 nucleotides, compared to 41 in alpha- and 54 in beta-globin mRNA. There is very little homology between alpha and gamma sequences in the 5' region. There is considerable homology between beta- and gamma-globin mRNAs in the regions proximal and distal to the initiation codon, but the entire sequence shows less homology than the human and rabbit beta-globin mRNAs. The hexanucleotide sequence CUUCUG is found near the 5' ends of all three human globin mRNAs, suggesting a possible role of this sequence or ribosomal binding. Both guanosine and cytidine were found at the 19th nucleotide position from the 5' end of the gamma mRNA. We believe this heterogeneity arises from the difference in nucleotide sequence between the A gamma and G gamma loci.
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PMID:The nucleotide sequence of the 5' untranslated region of human gamma-globin mRNA. 31 62

Sequence analysis studies were carried out on human beta-globin mRNA (beta-mRNA) prepared from alpha-thalassemic, sickle cell, and Hb A reticulocytes. Highly purified beta-mRNA served as substrate for the preparation of cDNA by RNA-dependent DNA polymerase. The cDNA was transcribed by Escherichia coli RNA polymerase and the resulting cRNA was analyzed. Over 300 nucleotides were assigned to the beta-mRNA coding region and 37 nucleotides were assigned to the 3'-terminal noncoding region. The normal termination codon is UAA which is separated by 28 nucleotides from an out of phase UAA triplet. The origin of each of the abnormally long beta-globin variants Tak and Cranston is consistent with reduplication of dinucleotides prior to the normal termination codon, and both globin variants can terminate at the out of phase UAA.
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PMID:Human beta-globin messenger RNA. I. Nucleotide sequences derived from complementary RNA. 87 28

Complementary DNA enriched in sequences hybridizing to beta-globin mRNA was prepared with viral RNA-dependent DNA polymerase and used as a probe for the presence of beta-globin mRNA in nuclear and cytoplasmic RNA from two Italian patients with beta0-thalassaemia. In both cases the beta-globin gene was present and cytoplasmic mRNAbeta was absent; however, one case appeared to transcribe mRNAbeta and to fail to process it, while the other appeared transcriptionally defective. Evidence is also presented that the low levels of hybridization usually found at high RNA/cDNAbeta ratios in beta0-thalassaemia are due to delta-globin mRNA; the melting profile of the hybrid formed has been determined and a low melting temperature relative to mRNAbeta - cDNAbeta demonstrated.
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PMID:Transcriptional and post-transcriptional defects in beta0-thalassaemia. 92 69

The specificity of hybridization was compared between the human and rabbit alpha and beta-globin complementary DNAs (cDNAs) and the corresponding alpha and beta-globin messenger RNAs (mRNAs). The globin chain-specific mRNAs of rabbit were prepared from polysomes incubated with O-methylthreonine (alpha and beta) or from postribosomal supernatant (alpha). Enrichment for either the alpha- or beta-globin mRNA was demonstrated by cell-free protein synthesis and by RNA-cDNA hybridization. Human mRNAs, active as templates for RNA-directed DNA polymerase, were prepared from reticulocytes of patients with hemolytic anemia, alpha-thalassemia (hemoglobin H disease), and beta-thalassemia. Because there was partial cross-hybridization between human mRNA and rabbit cDNA, the rabbit alpha- and beta-globin cDNAs could be used to demonstrate that the beta-thalassemia mRNA was enriched in human alpha-globin mRNA sequences and that the alpha-thalassemia mRNA was enriched in human beta-globin mRNA sequences. These results were confirmed by preparation of thalassemia globin cDNAs and subsequent hybridization to their template mRNAs. The amount of cross-hybridization between the human and rabbit alpha-globin mRNA and the two alpha-globin cDNAs was comparable to the cross-hybridization between the two beta-globin mRNAs and the two beta-globin cDNAs, indicating a similar degree of evolutionary divergence in the nucleotide sequences of the two globin genes.
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PMID:alpha-and beta-Globin complementary deoxyribonucleic acids of human and rabbit. Specificity of hybridization. 112 37


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