| 1. |
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| 2. |
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| 3. |
- Albinsson, Bo, 1963, et al.
(författare)
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The Electronically Excited-States of 2-Phenylindole
- 1991
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Ingår i: Chemical Physics. - 0301-0104. ; 151:1, s. 149-157
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Tidskriftsartikel (refereegranskat)abstract
- The light absorption of 2-phenylindole (2PI) in the UV region (210-350 nm) is investigated by means of linear dichroism in stretched polyethylene film and fluorescence polarization anisotropy in a propylene glycol glass. Experimentally, 2PI is found to have five distinct electronic transitions located above 200 nm for which the transition moments are determined. The conclusions are supported by quantum chemical calculations and the origin of the so-called composite band is discussed. Comparison with the absorption spectrum of the fluorescent DNA binding probe DAPI is also made and 2PI is found to be an appropriate model system for the electronic transitions of DAPI.
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| 4. |
- Bar, Tzachi, 1970, et al.
(författare)
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Kinetic Outlier Detection (KOD) in real-time PCR.
- 2003
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Ingår i: Nucleic acids research. - 1362-4962 .- 0305-1048. ; 31:17
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Tidskriftsartikel (refereegranskat)abstract
- Real-time PCR is becoming the method of choice for precise quantification of minute amounts of nucleic acids. For proper comparison of samples, almost all quantification methods assume similar PCR efficiencies in the exponential phase of the reaction. However, inhibition of PCR is common when working with biological samples and may invalidate the assumed similarity of PCR efficiencies. Here we present a statistical method, Kinetic Outlier Detection (KOD), to detect samples with dissimilar efficiencies. KOD is based on a comparison of PCR efficiency, estimated from the amplification curve of a test sample, with the mean PCR efficiency of samples in a training set. KOD is demonstrated and validated on samples with the same initial number of template molecules, where PCR is inhibited to various degrees by elevated concentrations of dNTP; and in detection of cDNA samples with an aberrant ratio of two genes. Translating the dissimilarity in efficiency to quantity, KOD identifies outliers that differ by 1.3-1.9-fold in their quantity from normal samples with a P-value of 0.05. This precision is higher than the minimal 2-fold difference in number of DNA molecules that real-time PCR usually aims to detect. Thus, KOD may be a useful tool for outlier detection in real-time PCR.
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| 5. |
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| 6. |
- Bengtsson, Martin, et al.
(författare)
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Gene expression profiling in single cells from the pancreatic islets of Langerhans reveals lognormal distribution of mRNA levels.
- 2005
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Ingår i: Genome Research. - : Cold Spring Harbor Laboratory. - 1549-5469 .- 1088-9051. ; 15:10, s. 1388-1392
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Tidskriftsartikel (refereegranskat)abstract
- The transcriptional machinery in individual cells is controlled by a relatively small number of molecules, which may result in stochastic behavior in gene activity. Because of technical limitations in current collection and recording methods, most gene expression measurements are carried out on populations of cells and therefore reflect average mRNA levels. The variability of the transcript levels between different cells remains undefined, although it may have profound effects on cellular activities. Here we have measured gene expression levels of the five genes ActB, Ins1, Ins2, Abcc8, and Kcnj11 in individual cells from mouse pancreatic islets. Whereas Ins1 and Ins2 expression show a strong cell-cell correlation, this is not the case for the other genes. We further found that the transcript levels of the different genes are lognormally distributed. Hence, the geometric mean of expression levels provides a better estimate of gene activity of the typical cell than does the arithmetic mean measured on a cell population.
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| 7. |
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| 8. |
- Eriksson, Svante, 1957, et al.
(författare)
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BINDING OF 4',6-DIAMIDINO-2-PHENYLINDOLE (DAPI) TO AT REGIONS OF DNA - EVIDENCE FOR AN ALLOSTERIC CONFORMATIONAL CHANGE
- 1993
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Ingår i: Biochemistry. - : American Chemical Society (ACS). - 1520-4995 .- 0006-2960. ; 32:12, s. 2987-2998
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Tidskriftsartikel (refereegranskat)abstract
- The interaction of 4',6-diamidino-2-phenylindole (DAPI) with several double-helical poly- and oligonucleotides has been studied in solution using optical spectroscopic techniques: flow linear dichroism (LD), induced circular dichroism (CD), and fluorescence spectroscopy. In AT-rich sequences, where DAPI is preferentially bound, LD indicates that the molecule is edgewise inserted into the minor groove at an angle of approximately 45-degrees to the helix axis. This binding geometry is found for very low as well as quite high binding ratios. The concluded geometry is in agreement with that of the DAPI complex in a crystal with the Drew-Dickerson dodecamer, and the DAPI complex with this dodecamer in solution is verified to have an ICD spectrum similar to that of the complex with [poly(dA-dT)]2 at low binding ratios. The observation of two types of CD spectra characteristic for the binding of DAPI to DNA, and also for the interaction with [poly(dA-dT)]2, demonstrates that the first binding mode, despite its low apparent abundance (a few percent), is not due to a specific DNA site. The effect may be explained in terms of an allosteric binding such that when DAPI molecules bind contiguously to the AT sequence the conformation of the latter is changed. The new conformation, which according to LD appears to be stiffer than normal B-form DNA, is responsible for the second type of induced CD spectrum in the DAPI chromophore. Although the spectroscopic results indicate a change of DNA conformation, consistent with an allosteric binding model, they do not explicitly require any cooperativity, but accidental neighbors could also explain the data.
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| 9. |
- Ghasemi, J., et al.
(författare)
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A new algorithm for the determination of protolytic constants from spectrophotometric data in multiwavelength mode : Calculations of acidity constants of 4-(2-pyridylazo)resorcinol (PAR) in mixed nonaqueous-water solvents
- 2006
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Ingår i: Talanta. - : Elsevier BV. - 0039-9140 .- 1873-3573. ; 68:4, s. 1201-1214
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Tidskriftsartikel (refereegranskat)abstract
- A new efficient, simple and versatile algorithm is presented for determination of the protolytic constants from spectrophotmetric data in multiwavelength mode based on the combining of hard and soft modeling. The algorithm was checked by determining the acidity constants of a triprotic acid from theoretical and real absorption-pH data. The real spectral data are obtained from photometric titration of different solutions of 4-(2-pyridylazo)resorcinol (PAR) by a standard base solution under an inert atmosphere. The algorithm starts the minimization process using an user supplied number of components and initial guesses of the unknown parameters and refined in a least squares manner. New algorithm is implemented in the new version of DATAN package (version 3.1). The validity of the obtained results was checked by some well known programs such as DATAN 2.1, SPECFIT/32, SQUAD, a modified version of difference spectra and a A-pH curve method. The comparison of the outputs of the DATAN 3.1 with the other programs reveals that there is a very good agreement between the obtained results and mentioned programs.
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| 10. |
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| 11. |
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| 12. |
- Hagmar, Per, et al.
(författare)
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IONIC-STRENGTH DEPENDENCE OF THE BINDING OF METHYLENE-BLUE TO CHROMATIN AND CALF THYMUS DNA
- 1992
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Ingår i: Journal of Biomolecular Structure and Dynamics. - 0739-1102 .- 1538-0254. ; 9:4, s. 667-679
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Tidskriftsartikel (refereegranskat)abstract
- The binding of the intercalating dye methylene blue (MB) to chromatin and to free DNA has been studied as a function of ionic strength at very low binding ratios (1 MB/400 DNA bases) using absorption spectroscopy. With increasing salt concentration MB is displaced from chromatin to a higher extent than from DNA. The free energy change for MB binding to chromatin is found to be approximately 5 kJ/mole lower than for binding to DNA. This difference can be explained by the reduced number of high affinity binding sites in chromatin due to the presence of histone proteins. The difference in binding energy is virtually independent of the degree of chromatin condensation and also of the valence of counter ions, suggesting that neither the affinity for, nor the number of intercalation sites in the linker DNA is markedly changed upon the salt-induced condensation. The unaffected thermodynamics of the linker binding suggests that factors such as DNA superhelicity and the electrostatic influence from the chromatosomes remain unchanged during chromatin condensation.
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| 13. |
- Håkansson, Joakim, 1975, et al.
(författare)
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Neural cell adhesion molecule-deficient beta-cell tumorigenesis results in diminished extracellular matrix molecule expression and tumour cell-matrix adhesion
- 2005
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Ingår i: Tumour Biology. - : Springer Science and Business Media LLC. - 1010-4283 .- 1423-0380. ; 26:2, s. 103-112
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Tidskriftsartikel (refereegranskat)abstract
- To understand by which mechanism neural cell adhesion molecule (N-CAM) limits beta tumour cell disaggregation and dissemination, we searched for potential downstream genes of N-CAM during beta tumour cell progression by gene expression profiling. Here, we show that N-CAM-deficient beta-cell tumorigenesis is associated with changes in the expression of genes involved in cell-matrix adhesion and cytoskeletal dynamics, biological processes known to affect the invasive and metastatic behaviour of tumour cells. The extracellular matrix (ECM) molecules emerged as the primary target, i.e. N-CAM deficiency resulted in down-regulated mRNA expression of a broad range of ECM molecules. Consistent with this result, deficient deposition of major ECM stromal components, such as fibronectin, laminin 1 and collagen IV, was observed. Moreover, N-CAM-deficient tumour cells displayed defective matrix adhesion. These results offer a potential mechanism for tumour cell disaggregation during N-CAM-deficient beta tumour cell progression. Prospective consequences of these findings for the role of N-CAM in beta tumour cell dissemination are discussed.
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| 14. |
- Jansen, Karl, 1966, et al.
(författare)
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SEQUENCE DEPENDENCE OF 4',6-DIAMIDINO-2-PHENYLINDOLE (DAPI) DNA INTERACTIONS
- 1993
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 1520-5126 .- 0002-7863. ; 115:23, s. 10527-10530
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Tidskriftsartikel (refereegranskat)abstract
- We have characterized the interaction of 4',6-diamidino-2-phenylindole (DAPI) with a series of DNA oligonucleotides containing a varying number of contiguous alternating AT base pairs in the center of a GC-stretch. The binding affinity of DAPI to the oligonucleotides increases with the number of contiguous AT base pairs. When entropy effects are taken into account, the microscopic affinity reaches a maximum value when the oligonucleotides contain three AT base pairs. Three is also the minimum number of base pairs that provides a binding site with all the characteristics of binding observed with DNA and [poly(dA-dT)]2 at low binding ratios. With none or only a single AT base pair in the center of a GC dodecamer, the spectral properties of bound DAPI are very similar to those of DAPI bound to [poly(dG-dC)]2. When DAPI is complexed with an oligonucleotide containing two AT base pairs, the absorption and fluorescence properties of DAPI closely resemble those of DAPI complexed with [poly(dG-dC)]2, while the circular dichroism spectrum resembles that of DAPI bound to [poly(dA-dT)]2.
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| 15. |
- Kim, Seog K., et al.
(författare)
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INTERACTION OF 4',6-DIAMIDINO-2-PHENYLINDOLE (DAPI) WITH POLY D(G-C)2 AND POLY D(G-M(5)C)2 - EVIDENCE FOR MAJOR GROOVE BINDING OF A DNA PROBE
- 1993
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Ingår i: Journal of the American Chemical Society. - : American Chemical Society (ACS). - 1520-5126 .- 0002-7863. ; 115:9, s. 3441-3447
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Tidskriftsartikel (refereegranskat)abstract
- The interactions of DAPI with poly[d(G-C)2] and poly[d(G-m5C)2] are characterized by optical spectroscopic methods. DAPI is found to bind to poly[d(G-C)2] with its molecular long-axis roughly perpendicular but with its plane more parallel to the belix axis of the polynucleotide, probably being bound in a non-intercalated way in the major groove of the polynucleotide. In poly[d(G-m5C)2], DAPI is bound with its molecular plane perpendicular to the helix axis, probably partially intercalated from the major groove. The conclusions are supported by fluorescence quenching results. Together with the earlier established minor groove binding mode of DAPI in the AT regions, our results demonstrate a heterogeneous nature of the interaction of a small ligand with DNA. We conclude that the conventional classification of a ligand as being either intercalator, minor groove binder, or major groove binder may not be adequate since the mode of binding clearly depends also on the base sequence.
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| 16. |
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| 17. |
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| 18. |
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| 19. |
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| 20. |
- Kubista, Mikael, 1961, et al.
(författare)
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Determination of Equilibrium-Constants by Chemometric Analysis of Spectroscopic Data
- 1993
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Ingår i: Analytical Chemistry. - 0003-2700 .- 1520-6882. ; 65:8, s. 994-998
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Tidskriftsartikel (refereegranskat)abstract
- We describe a chemometric method to determine equilibrium constants with high accuracy from spectroscopic titrations. Knowledge of component spectra is not required for the analysis, and titrations can be analyzed even when the titration end points are not reached. In fact, the component spectra are determined in the analysis. The analysis is based on a decomposition of the recorded spectra into a product of target and projection matrices using NIPALS. The matrices are then rotated to give the correct concentrations and spectral profiles of the components utilizing the functional form of the equilibrium expression. The output of the analysis is the equilibrium constant and the component spectral profiles. The analysis is highly accurate and can be performed on a standard personal computer. As examples, we determine the protolytic constant for the equilibrium between the anionic and dianionic forms of fluorescein in aqueous solution and the dimerization constant of benzoic acid in n-heptane.
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| 21. |
- Kubista, Mikael, 1961, et al.
(författare)
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DNA diagnostics gets digitised
- 2011
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Ingår i: Drug Discovery World. ; :Fall, s. 77-82
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Quantitative real-time PCR (qPCR) has during the last two decades emerged as the preffered technology for nucleic acid analysis in routine as well as in research. qPCR has the sensitivity to detect a single molecule, the specificity to differentiate targets by a single nucleotide, and, because of its exponential nature, virtually unlimitied dynamic range
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| 22. |
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| 23. |
- Kubista, Mikael, 1961, et al.
(författare)
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High Troughput single cell expression profiling: taking a closer look on biological
- 2011
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Ingår i: European Pharmaceutical Review. ; 16:2, s. 20-24
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Molecular analysis of tissue and in most cases also of bodily fluids is complicated because of tissue heterogeneity and the presence of many different cell types. Even cells of apparently the same type show substantial variation in gene expression under virtually identical conditions. When analysing classical samples based on tens of thousands of cells, this natural variability among cells is lost. With the advent of real-time quantitative polymerase chain reaction (qPCR), we have a most powerful tool to study diversity on the single cell level and can detect rare cells that are critical to treatment or survival.
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