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Tony Minson - One of the best experts on this subject based on the ideXlab platform.

Andalfred I Geller - One of the best experts on this subject based on the ideXlab platform.

  • helper virus free transfer of herpes simplex virus type 1 Plasmid Vectors into neural cells
    Journal of Virology, 1996
    Co-Authors: Cornel Fraefel, Song Song, Filip Lim, Phung Lang, Yaming Wang, Peter J Wild, Andalfred I Geller
    Abstract:

    Herpes simplex virus type 1 (HSV-1) Plasmid Vectors have promise for genetic intervention in the brain, but several problems caused by the helper virus have compromised their utility. To develop a helper virus-free packaging system for these Vectors, the DNA cleavage/packaging signals were deleted from a set of cosmids that represents the HSV-1 genome. Following cotransfection into cells, this modified cosmid set supported replication and packaging of vector DNA. However, in the absence of the DNA cleavage/packaging signals, the HSV-1 genome was not packaged, and consequently vector stocks were free of detectable helper virus. In the absence of helper virus, the Vectors efficiently infected rat neural cells in culture or in the brain with minimal cytopathic effects. beta-galactosidase-positive cells were observed for at least 1 month in vivo, and vector DNA persisted for this period. This system may facilitate studies on neuronal physiology and potential therapeutic applications.

Helena Browne - One of the best experts on this subject based on the ideXlab platform.

Barbara E.h. Coupar - One of the best experts on this subject based on the ideXlab platform.

Joseph Sambrook - One of the best experts on this subject based on the ideXlab platform.

  • Preparation of Plasmid DNA by Alkaline Lysis with Sodium Dodecyl Sulfate: Maxipreps.
    Cold Spring Harbor protocols, 2018
    Co-Authors: Michael R Green, Joseph Sambrook
    Abstract:

    In recent years, with the advent of polymerase chain reaction and the development of highly efficient methods of cloning and DNA sequencing, the need to prepare large quantities of Plasmid Vectors and recombinants has greatly diminished. In consequence, this protocol, at one time in common use, has been largely replaced by faster and easier column-based purification methods. In this protocol, Plasmid DNA is purified from the cleared bacterial lysate by centrifugation to equilibrium in CsCl gradients containing ethidium bromide. These gradients have an aesthetic quality that justifies the continued presentation of a protocol that in many other respects is an antique. The typical yield of high-copy-number Plasmid Vectors or of amplified low-copy-number Vectors prepared by this method is ∼3-5 µg of DNA/mL of original bacterial culture. The yield of recombinant Plasmids containing inserts of foreign DNA is usually slightly lower, depending on the size and nature of the cloned DNA fragment.

  • the condensed protocols from molecular cloning a laboratory manual
    2006
    Co-Authors: Joseph Sambrook, David W Russell
    Abstract:

    1. Using Plasmid Vectors in Molecular Cloning 2. Bacteriophage bb and Its Vectors 3. Working with Bacteriophage M13 Vectors 4. Working with High-capacity Vectors 5. Gel Electrophoresis of DNA and Pulsed-field Agarose Gel Electrophoresis 6. Preparation and Analysis of Eukaryotic Genomic DNA 7. Extraction, Purification, and Analysis of mRNA from Eukaryotic Cells 8. In Vitro Amplification of DNA by the Polymerase Chain Reaction 9. Preparation of Radiolabeled DNA and RNA Probes 10. Working with Synthetic Oligonucleotide Probes 11. Preparation of cDNA Libraries and Gene Identification 12. DNA Sequencing 13. Mutagenesis 14. Screening Expression Libraries 15. Expression of Cloned Genes in Escherichia coli 16. Introducing Cloned Genes into Cultured Mammalian Cells 17. Analysis of Gene Expression in Mammalian Cells 18. Protein Interaction Technologies Appendices Index