Ultrafast DNA Research Articles

Imaging of a band for DNA fragment migrating in microchannel on integrated microchip

We investigated the microfabrication of PCR reaction vessel, capillary array electrophoresis (CAE), and laser-induced fluorescence detection cell using synchrotron radiation in polymethylmethacrylate (PMMA) substrates towards the development of novel technology for ultra-fast DNA analysis. Laser-induced fluorescence detection system for the chip has been developed and successfully applied to the imaging of a band for DNA fragment labeled by fluorescent dye during migrating in the array of microchannels to optimize the separation conditions for DNA analysis. Based on the imaging data, we can estimate that 10-s separation of some DNA fragments will be achievable using 10-mm separation channel on a chip.

Ultrafast DNA analysis by capillary electrophoresis/laser-induced fluorescence detection.

The limits of ultrafast DNA analysis by CE were determined by investigating the influence of the effective capillary length and the electric field strength on the analysis time for a given peak resolution (10 bp). In accordance with theory, the use of a fast ramp power supply for narrow plug electrokinetic injection was found to be essential to minimize the extra column effects on peak dispersion. Two major column dispersion factors, longitudinal diffusion and thermal dispersion, were determined experimentally, as well as the influence of the electric field strength on the electrophoretic mobilities and diffusion coefficients of DNA. It was found that higher field strengths can be applied with lower thermal dispersion than predicted by classical CE models. This was attributed to the faster mass transport in the radial direction due to field-induced DNA orientation. Short capillaries (approximately 3-7 cm effective length) and moderate to high electric field strengths (approximately 600-800 V/cm) were used to perform a series of fast DNA separations. The dsDNA fragment standards phiX174/HaeIII and pBR322/HaeIII were separated within 30 s. The possibility for fast mutation detection was demonstrated using constant denaturant capillary electrophoresis (CDCE) for the analysis of a single base mutation in mitochondrial DNA in 72 s. The potential for fast DNA sequencing was illustrated by separating 300 ssDNA fragments within 180 s.

Use of the restriction enzyme AvaI and exo- Bst polymerase in strand displacement amplification.

BioTechniquesVol. 24, No. 3 BenchmarksOpen AccessUse of the Restriction Enzyme AvaI and Exo− Bst Polymerase in Strand Displacement AmplificationMaria A. Milla, Patricia A. Spears, Robert E. Pearson & G. Terrance WalkerMaria A. Milla*Address correspondence to G. Terrance Walker or Maria A. Milla, Department of Molecular Biology, Becton Dickinson Research Center, PO Box 12016, 21 Davis Drive, Research Triangle Park, NC 27709-2016, USA. Internet: E-mail Address: walker@bdrc.bd.com or E-mail Address: milla@bdrc.bd.comBecton Dickinson Research Center, Research Triangle Park, NC, USASearch for more papers by this author, Patricia A. SpearsBecton Dickinson Research Center, Research Triangle Park, NC, USASearch for more papers by this author, Robert E. PearsonBecton Dickinson Research Center, Research Triangle Park, NC, USASearch for more papers by this author & G. Terrance Walker*Address correspondence to G. Terrance Walker or Maria A. Milla, Department of Molecular Biology, Becton Dickinson Research Center, PO Box 12016, 21 Davis Drive, Research Triangle Park, NC 27709-2016, USA. Internet: E-mail Address: walker@bdrc.bd.com or E-mail Address: milla@bdrc.bd.comBecton Dickinson Research Center, Research Triangle Park, NC, USASearch for more papers by this authorPublished Online:15 Aug 2018https://doi.org/10.2144/98243bm12AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinkedInRedditEmail FiguresReferencesRelatedDetailsCited ByRapid, Visual, and Sequence-Specific Detection of Salmonella in Egg Liquid with vis-NEAA, a CRISPR/Cas12 Empowered New Strategy9 February 2022 | Journal of Agricultural and Food Chemistry, Vol. 70, No. 7Strand displacement DNA synthesis by DNA polymerase gp90 exo― of Pseudomonas aeruginosa phage 1Biochimie, Vol. 170Photochemical Acceleration of DNA Strand Displacement by Using Ultrafast DNA Photo-crosslinking29 August 2017 | ChemBioChem, Vol. 18, No. 20Regulation of yeast DNA polymerase δ-mediated strand displacement synthesis by 5′-flaps26 March 2015 | Nucleic Acids Research, Vol. 43, No. 8Optimization and design of oligonucleotide setup for strand displacement amplificationJournal of Biochemical and Biophysical Methods, Vol. 63, No. 3Strand displacement amplification: a versatile tool for molecular diagnostics9 January 2014 | Expert Review of Molecular Diagnostics, Vol. 4, No. 2DNA Polymerases as Engines for BiotechnologyScott C. Hamilton, Joseph W. Farchaus & Maria C. Davis5 September 2018 | BioTechniques, Vol. 31, No. 2SDA Target AmplificationReal-Time, Sequence-Specific Detection of Nucleic Acids during Strand Displacement AmplificationAnalytical Biochemistry, Vol. 276, No. 2Characterization of an Extremely Thermostable Restriction Enzyme, Psp GI, from a Pyrococcus Strain and Cloning of the Psp GI Restriction-Modification System in Escherichia coliApplied and Environmental Microbiology, Vol. 64, No. 10Non–Polymerase Chain Reaction Mediated Target Amplification Techniques Vol. 24, No. 3 Follow us on social media for the latest updates Metrics Downloaded 383 times History Published online 15 August 2018 Published in print March 1998 Information© 2018 Author(s)PDF download

DNA in Non Hodgkin-Lymphoma —A Cytophotometric Study

Cytophotometric DNA determinations on 26 cases of non-Hodgkin's lymphoma yielded the following findings: 1. Follicular centrocytic/centroblastic lymphomas (M. Brill-Symmers) and diffuse centrocytic lymphomas (lymphocytic lymphosarcoma) have a diploid DNA stem line. Diploid DNA values are observed in benign tumors, so that the assignment of these lymphomas to the group of "low grade malignancies" appears justified. 2. Lymphoblastic sarcomas show an aneuploid DNA stem line, as do 96% of all malignant tumors. 3. Lymphoid cells, plasma cells, and immunoblasts seen in immunocytomas are aneuploid. Thus these lymphomas must belong to the group of "high-grade malignant lymphomas" as regards their DNA distribution. 4. Immunoblastic sarcomas have aneuploid DNA stem lines (1 case tetraploid), in which both the lymphoid cells and the plasma cells from those immunoblastic sarcomas arising from immunocytomas show atypical DNTA distribution patterns. 5. In two cases of angioimmunoblastic lymphadenopathy, the lymphoid cells, plasma cells, and immunoblasts are aneuploid. They are thus regarded as "high grade malignancy" lymphomas. The results are discussed with respect to clinical course and prognosis. Measurements on a larger series of cases and correlation to clinical data are needed to support these results. Ultrafast DNA measurements made by flow-through cytophotometry can perhaps be helpful in the future for making the decision between a "low" or "high" grade malignant lymphoma.