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- Glemarec, C, et al.
(författare)
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The NMR structure of 31-mer RNA domain of E. coli RNase P RNA using its non-uniformly deuterium labelled counterpart (the "NMR-window" concept)
- 1996
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Ingår i: Nucleic Acids Research. - : Oxford University Press (OUP). - 0305-1048 .- 1362-4962. ; 24:11, s. 2022-2035
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Tidskriftsartikel (refereegranskat)abstract
- The NMR structure of a 31mer RNA constituting a functionally important domain of the catalytic RNase P RNA from Escherichia coli is reported. Severe spectral overlaps of the proton resonances in the natural 31mer RNA (1) were successfully tackled by unique spectral simplifications found in the partially-deuterated 31 mer RNA analogue (2) incorporating deuterated cytidines [C5 (>95 atom % 2H), C2' (>97 atom % 2H), C3' (>97 atom % 2H), C4' (>65 atom % 2H) and C5' (>97 atom % 2H)] [for the 'NMR-window' concept see: Földesi,A. et al. (1992) Tetrahedron, 48, 9033; Foldesi,A. et al. (1993) J. Biochem. Biophys. Methods, 26, 1; Yamakage,S.-I. et al. (1993) Nucleic Acids Res., 21, 5005; Agback,P. et al. (1994) Nucleic Acids Res., 22, 1404; Földesi,A. et al. (1995) Tetrahedron, 51, 10065; Földesi,A. et al. (1996) Nucleic Acids Res., 24, 1187-1194]. 175 resonances have been assigned out of total of 235 non-exchangeable proton resonances in (1) in an unprecedented manner in the absence of 13C and 15N labelling. 41 out of 175 assigned resonances could be accomplished with the help of the deuterated analogue (2). The two stems in 31mer RNA adopt an A-type RNA conformation and the base-stacking continues from stem I into the beginning of the loop I. Long distance cross-strand NOEs showed a structured conformation at the junction between stem I and loop I. The loop I-stem II junction is less ordered and shows structural perturbation at and around the G11 -C22 base pair.
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- Hultdin, Magnus, et al.
(författare)
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Telomere analysis by fluorescence in situ hybridization and flow cytometry
- 1998
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Ingår i: Nucleic Acids Research. - Oxford : Oxford University Press. - 0305-1048 .- 1362-4962. ; 26:16, s. 3651-3656
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Tidskriftsartikel (refereegranskat)abstract
- Determination of telomere length is traditionally performed by Southern blotting and densitometry, giving a mean telomere restriction fragment (TRF) value for the total cell population studied. Fluorescence in situ hybridization (FISH) of telomere repeats has been used to calculate telomere length, a method called quantitative (Q)-FISH, We here present a quantitative flow cytometric approach, Q-FISHFCM, for evaluation of telomere length distribution in individual cells based on in situ hybridization using a fluorescein-labeled peptide nucleic acid (PNA) (CCCTAA)(3) probe and DMA staining with propidium iodide, A simple and rapid protocol with results within 30 h was developed giving high reproducibility, One important feature of the protocol was the use of an internal cell line control, giving an automatic compensation for potential differences in the hybridization steps. This protocol was tested successfully on cell lines and clinical samples from bone marrow, blood, lymph nodes and tonsils. A significant correlation was found between Southern blotting and Q-FISHFCM telomere length values (P = 0.002), The mean sub-telomeric DNA length of the tested cell lines and clinical samples was estimated to be 3.2 kbp, With the Q-FISHFCM method the fluorescence signal could be determined in different cell cycle phases, indicating that in human cells the vast majority of telomeric DNA is replicated early in S phase.
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- Johanson, Urban, et al.
(författare)
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A new mutation in 16S rRNA of Escherichia coli conferring spectinomycin resistance
- 1995
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Ingår i: Nucleic Acids Research. - 0305-1048 .- 1362-4962. ; 23:3, s. 464-466
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Tidskriftsartikel (refereegranskat)abstract
- We report a novel mutation, Cl 066U in 16S rRNA whichwas selected for resistance to spectinomycin, anantibiotic which inhibits ribosomal translocation. Theminimal inhibitory concentration (MIC) of spectinomycindetermined for this mutant (15 pg/ml) is greaterthan with the wild-type plasmid (5 ig/ml) but lower thanwith the well known C1192U mutation (>80 pg/ml). TheCl 066U mutation also increases the cells sensitivity tofusidic acid, another antibiotic which inhibits translationat the translocation stage, whereas C1192U isunchanged relative to the wild type. We discuss whythe acquisition of resistance to one of these drugs isoften associated with hypersensitivity to the other.
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