By Russell J. Diefenbach, Cornel Fraefel
Herpes Simplex Virus: tools and Protocols presents a large choice of protocols hired in a variety of degrees of herpes virus study, together with easy protocols on growing to be viruses in cellphone tradition and cloning, manipulating and getting ready viral DNA. different chapters describe ways to layout and practice HSV-1 vectors for vaccination, melanoma and gene treatment or to review particular facets of HSV-1 biology resembling latency, intracellular delivery and protein-protein interplay. strategies for structural analyses, microscopy, proteomics and trying out of antivirals are incorporated besides. Written within the hugely winning Methods in Molecular Biology sequence structure, chapters comprise introductions to their respective subject matters, lists of the required fabrics and reagents, step by step, with ease reproducible laboratory protocols and pointers on troubleshooting and fending off recognized pitfalls.
Practical and authoritative, Herpes Simplex Virus: equipment and Protocols will relief new researchers within the box of herpes virology in addition to these skilled investigators wishing to embark on new techniques.
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Herpes Simplex Virus: equipment and Protocols offers a large choice of protocols hired in a number of degrees of herpes virus study, together with easy protocols on becoming viruses in telephone tradition and cloning, manipulating and getting ready viral DNA. different chapters describe methods to layout and practice HSV-1 vectors for vaccination, melanoma and gene treatment or to review particular elements of HSV-1 biology corresponding to latency, intracellular shipping and protein-protein interplay.
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Additional info for Herpes Simplex Virus: Methods and Protocols
19. HSV viral stock solution, titered to determine the number of plaque-forming units (pfu) per milliliter. 1 Methods Day 1 1. Check that host cell monolayers are confluent. Trypsinize one plate, and count cells using a hemocytometer. Calculate the number of cells per dish, based on dilution used for hemocytometer counting. 2. , if each dish has 1 × 106 cells, then add 5 × 106 pfu per dish (see Note 6). 2 Day 2 1. Check for uniform cytopathic effect (CPE). Cells should be rounded but not floating (see Note 7).
PLoS One 8:e57224 4. Vahlne AG, Blomberg J (1974) Purification of herpes simplex virus. J Gen Virol 22:297–302 5. Knop DR, Harrell H (2007) Bioreactor production of recombinant herpes simplex virus vectors. Biotechnol Prog 23:715–721 6. Jiang C, Wechuck JB, Goins WF, Krisky DM, Wolfe D, Ataai MM, Glorioso JC (2004) Immobilized cobalt affinity chromatography provides a novel, efficient method for herpes simplex virus type 1 gene vector purification. J Virol 78:8994–9006 Chapter 3 Isolation of Herpes Simplex Virus Nucleocapsid DNA Moriah Szpara Abstract As an inanimate virus, herpes simplex virus type 1 (HSV-1) necessarily encodes all of its functions in its DNA.
6. 15 and 50 ml centrifuge screw cap tubes. 7. 50-ml tubes (Nalgene Centrifuge Oak Ridge copolymer). 8. 2 ml capacity (Beckman, prod. No 362181). 9. Needles: 18 G, 1½ in. 10. Syringes: 10 cc. 11. Sonicator Ultrasonic Processor with 2½″ Cup Horn. 12. Centrifuge Beckman Avanti J25 with JA-20 rotor. 13. 1 rotor. 14. Iodixanol (Optiprep™, Axis-Shield, prod. No 1030061 Norway). 15. 0. 22 μm. 22 Peggy Marconi and Roberto Manservigi Store at 4 °C. 0. Add H2O to a final volume of 100 ml and filter sterilize as solution B.