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1.
Previous work has indicated that loss of ability of DNA dependent RNA polymerase, from stationary phase cultures of B. subtilis, to transcribe phage øe DNA was a sine qua non for sporulation. To ascertain if this change in template specificity was sporulation-specific, we repeated these experiments using a defined sporulation medium. The changes observed previously did not occur in the defined medium although sporulation was normal. The ability of the enzyme to transcribe other DNA templates was also examined. Similar studies were carried out using a polymerase from a rifamycin-resistant, sporulation conditional mutant. The significance of these findings with regard to the regulation of sporulation in B. subtilis is discussed.  相似文献   

2.
An endonuclease has been isolated and purified from Escherichiacoli which degrades RNA hydrogen bonded to DNA and no other polynucleotide substrates, including double stranded RNA, single stranded RNA, double stranded DNA or single stranded DNA.  相似文献   

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Reiteration of DNA complementary to a cytoplasmic non-ribosomal RNA   总被引:1,自引:0,他引:1  
Experimentally induced granulomas, in guinea pigs, were fractionated into a 40,000xg pellet and supernatant, which was further fractionated into a 100,000xg pellet and supernatant. The product of in vitro amino acid incorporation by the 40,000xg pellet was tentatively identified as collagen by its high proline/leucine ratio, its digestibility by bacterial collagenase and its solubility in hot trichloroacetic acid. The 100,000xg pellet incorporated leucine much more efficiently than the 40,000xg pellet and the product was insoluble in hot trichloroacetic acid.Labeled RNA from the 40,000xg pellet formed hybrids with granuloma (Cot 12 = 100–150) and liver (Cot 12 = 8,000) chromatin DNA, indicating that genes coding for this RNA are repeated about 100-fold in granuloma and less than 5 times in liver DNA. Under conditions of poly(A) binding, 50% of this labeled RNA is retained by filters. Digestion with ribonuclease and ribonuclease T1, decreases binding efficiency by 75%.  相似文献   

5.
The invitro DNA dependent synthesis of ribosomal protein L12 and the β subunit of RNA polymerase has been investigated using DNA from a plasmid which contains the genetic information for ribosomal protein L12 and the β subunit of RNA polymerase. This DNA, however, lacks the promoter region and the genetic information for the first 26 amino acids of ribosomal protein L10. It was found that L12 and the β subunit of RNA polymerase are efficiently synthesized invitro from this DNA. These results suggest that L12 and the β subunit of RNA polymerase can be synthesized from a promoter situated within the L10 gene.  相似文献   

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The equivalence of messenger RNA released (transported) from isolated rat liver nuclei to three selected media, with messenger RNA normally released to liver cytoplasm in vivo, has been evaluated by competitive DNA: RNA hybridization. Near normal nuclear restriction was exhibited by nuclei in media fortified with ATP, salts, spermidine and dialyzed cytosol. The RNA transport in the latter system was markedly inhibited by colchicine as was also the transport of RNA in vivo. Both nuclear restriction and sensitivity of the RNA transport to colchicine in media lacking spermidine and cytosol deviated significantly from the in vivo norm. The results emphasize the importance of establishing the in vivo equivalence in cell-free systems designed to study RNA synthesis, processing and transport.  相似文献   

8.
Replication of repeated DNA in human cells   总被引:1,自引:0,他引:1  
The replication pattern of the repeated sequence families of human DNA has been studied by means of DNA reassociation curves. Early- and late-replicating DNA fractions were obtained from synchronized cultures of KB cells by labeling cells with bromodeoxyuridine (BUdR) early or late in the DNA synthesis period and isolating the BUdR-containing DNA by CsCl density-gradient centrifugation. Highly repeated and moderately repeated sequence classes labeled with 14C-deoxycytidine either early or late in the DNA synthesis period were also prepared. The effect of the isolated early- or late-replicating BUdR-DNA on the rate of reassociation of the 14C-labeled repeated sequences was then tested. Increasing concentrations of early- or late-replicating BUdR-DNA were added to a constant amount of either 14C-labeled early- or late-replicating repeated sequences, and the fraction of label in double-stranded DNA was determined. Analysis of the DNA reassociation curves so obtained indicates that some repeated sequence families are replicated throughout the DNA synthesis period whereas others are replicated primarily in the second half. This is true for both the highly-repeated and moderately-repeated sequence classes.  相似文献   

9.
Clones of hybrid plasmids containing moderately and highly repeated sequences of Drosophila hydei exhibit positive autoradiographic signals if hybridized to labeled whole genome DNA. Such clones were screened with labeled male (XY) and female (XX) DNA, and male-specific fragments were identified. Further hybridization of male-specific clones to female XXY, and male X0 DNAs established them as containing Y-specific moderately repeated sequences. Further verification of one particular cloned fragment as Y-specific is presented and possible applications of this procedure are briefly discussed.  相似文献   

10.
Neomycin inhibits in vitro DNA dependent DNA and RNA synthesis catalyzed by DNA polymerase I and RNA polymerase from E. coli. The effect of the antibiotic is more pronounced towards DNA synthesis. The inhibition of DNA synthesis is competitive with template DNA, does not reverse with excess deoxynucleoside triphosphate, Mg2+ or enzyme E. coli DNA polymerase I. Neomycin does not reduce the number of potential 3′ -OH end or primer. It seems to shorten the size of the newly formed polynucleotide.  相似文献   

11.
Altered restriction of nuclear RNA during incubation in vitro   总被引:1,自引:0,他引:1  
Nuclei were isolated from rat liver and incubated invitro in two commonly employed RNA transport assays. Released [14C] RNA was isolated and hybridized with filter-bound DNA in the presence of competing cytoplasmic RNA. A significant portion of RNA which was transported to either medium was not represented in cytoplasmic RNA. These results indicate that the restriction of some sequences to the nucleus invivo is not maintained invitro.  相似文献   

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Purified RNA polymerase, DNA polymerase III and unwinding protein of Escherichiacoli catalyze limited rifampicin sensitive fd or ØX 174 DNA-dependent DNA synthesis. A protein has been partially purified from E.coli which stimulates rifampicin sensitive dXMP incorporation in this system 20 to 30 fold. This protein also stimulates DNA synthesis catalyzed by DNA polymerases I and II; the stimulation occurs in reactions primed with natural and synthetic DNAs as well as RNA-DNA hybrids. The protein is not a product of the known dna genes. In contrast to the above system of purified enzymes, rifampicin sensitive dXMP incorporation in crude extracts of E.coli is specifically dependent on fd but not ØX 174 DNA. An additional factor has been isolated from extracts of E.coli which restores specificity to the purified rifampicin sensitive system by preventing ØX 174 DNA from serving as a template.  相似文献   

14.
Conditions for the production of a complementary DNA sequence for use in studies of ribosomal RNA are described. E. coli DNA polymerase I is used to transcribe highly purified 28S ribosomal RNA from rat liver. The reaction is sensitive to the tertiary structure of the rRNA template-primer. The complementary DNA hybridizes to its rRNA template with a Rot12 of 0.02. The hybrid formed between 28S ribosomal RNA and complementary DNA has a Tm of 73°C. The probe reacts with total rat nuclear RNA with a Rot12 of 1.0.  相似文献   

15.
Using the plasmid pNF1337 as template, a mRNA preparation has been obtained that directs the in, vitro synthesis of fMet-Val, the N-terminal dipeptide of the β subunit of RNA polymerase. RNA polymerase holoenzyme specifically inhibits the mRNA-directed synthesis of fMet-Val showing that the autoregulation by RNA polymerase of β,β′ synthesis is at the level of translation. L factor (nusA gene product) stimulates the synthesis of fMet-Val from a DNA template but not from mRNA. Rifampicin has no effect on the mRNA-directed synthesis of fMet-Val or the ability of RNA polymerase to inhibit fMet-Val synthesis.  相似文献   

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Treatment of the eukaryotic organism Tetrahymena pyriformis with low concentrations of Ethidium Bromide causes accumulation of a protein-nucleic acid complex consisting of a DNA polymerase, a RNA polymerase, a deoxyribo-nuclease and a RNA linked DNA fragment. The length of the RNA is about 30 nucleotides, while the DNA part is around 200 nucleotides long. Degradations with ribonucleases and deoxyribonucleases strongly indicate that the RNA exists in a non-hybrid structure with a homogenous base composition and that the DNA is single-stranded. The complex is purified 1100 fold from whole cells and sodium dodecyl sulphate acrylamide gel electrophoresis gives 9 defined bands. The polynucleotide in the isolated complex accounts for only 10?4 of the total cellular DNA.As the complex contains some of the enzymes essential for discontinous DNA replication, in addition to a RNA linked Okazaki fragment, it is concluded that a highly purified replication complex has been isolated.  相似文献   

18.
A polypeptide isolated from red kidney beans, Phaseolusvulgaris, which has previously been shown to stimulate RNA synthesis in cultures of mouse spleen lymphocytes and plasmolyzed E.coli, is here shown to be a potent inhibitor of trypsin and α-chymotrypsin. This polypeptide is compared with commercially available trypsin inhibitors with regard to their capacity to inhibit some proteolytic enzymes and to stimulate invitro cultures of lymphocytes. Similar to FV the lima been trypsin inhibitor was found to possess a stimulating effect on the RNA as well as the DNA synthesis in lymphocyte cultures.  相似文献   

19.
An improved method has been developed for the isolation of nuclei from Saccharomyces cerevisiae for the study of RNA synthesis in vitro. Utilization of Ficoll in the isolation procedure greatly increases the activity of RNA polymerase in isolated nuclei. Nuclei prepared by this procedure are essentially free of mitochondrial DNA.  相似文献   

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