New PDF release: PCR Protocols: Current Methods and Applications
By Beverly C. Delidow, John P. Lynch, John J. Peluso, Bruce A. White (auth.), Bruce A. White (eds.)
The latest and most recent assortment, PCR Protocols bargains certain laboratory techniques for using polymerase chain reactions in a variety of functions. even though particular examples and experimental structures are defined, the book's concentration is on normal functions that may be changed to fit a vast spectrum of analysis platforms, therefore making its scope beautiful to almost all researchers. PCR Protocols beneficial properties transparent, easy-to-follow descriptions of tactics and a Notes part in every one bankruptcy to supply tips, substitute feedback, and different improvements of the protocols. Key issues and methods: easy startup approaches for PCR • PCR in detecting DNA and RNA • PCR in DNA synthesis and mutagenesis • PCR cloning of DNA • choice of primers • radioactive and nonradioactive labeling • quantitation of PCR items by means of HPLC • chromosome task • PCR mapping of HLA genes • sequencing of PCR items • genomic footprinting • quantitation of tumor gene expression • screening of phage libraries • SSP-PCR and genome jogging • subcloning of PCR items.
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Additional info for PCR Protocols: Current Methods and Applications
Southern Blotting and Hybridization 1. Agarose:SeaPlaqueor NuSieveGTG (PMC BioProducts,Rockland,ME). 2. Electrophoresisunit for submarinegelsanda constantvoltagepowersupply (Holzel, Dorfen, Germany). 3. Gel trays (11 x 13 cm) and a 1Cteeth comb bridge (Holzel, Dorfen, Germany). 4. Nylon membrane(11 x 13cm) (BoehringerMannheimGmbH, Mannheim, Germany). 5. , Maidstone,England). 6. Glass tray and glass plates or alternatively a vacuum-blotting device (VakuGene,PharmaciaP-L Biochemicals, Milwaukee, WI).
05% xylene cyan01 FF. 7. DNA sequencing apparatus (including glass plates, spacers, and combs). 3. Methods 1. Digest the 5’ ends of the PCR product by mixing 2 pL of PCR product (from a 100 uL reaction), 2 pL of 10X Klenow buffer, 1 pL of Klenow fragment, and 15 PL of H20 in a microfuge tube. 2. Let stand for 10 min at room temperature. 3. 5 pL of [a-3*]dCTP and 1 pL of dNTP solution. 4. Let stand 10 min at room temperature (see Note 1). 5. Destroy the enzyme by incubating at 70°C for 5 min. 6.
And Collins, F. S. (1991) Construction of T-vectors, a rapid and generalsystemfor direct cloning of unmodified PCR products Nucleic Acids Res. 19, 1154. 15. Holton, T. A. and Graham, M. W. (1991) A simple and efficient method for direct cloning of PCR products using ddT-tailed vectors Nuclex Acrds Res. 19, 1156. CHAPTER 3 Direct Radioactive Labeling of Polymerase Chain Reaction Products Tim McDaniel and Stephen J. Meltzer 1. Introduction Radioactively labeled polymerase chain reaction (PCR) products are being used in an increasing number of molecular biology research techniques.
PCR Protocols: Current Methods and Applications by Beverly C. Delidow, John P. Lynch, John J. Peluso, Bruce A. White (auth.), Bruce A. White (eds.)