This chapter reviews alternate methods for the generation of sequencing templates from amplified DNA and sequencing by the method of Sanger. Amplification of the cloned inserts of unknown sequence can be achieved using oligonucleotides that are priming inside, or close to, the polylinker of the cloning vector. In cases in which the optimization of polymerase chain reaction (PCR) conditions fail to produce the desired priming specificity, either new oligonucleotides are required or the different PCR products can be separated by gel electrophoresis and re-amplified individually for sequencing. The PCR method for the in vitro amplification of specific DNA fragments has opened up a number of fields in molecular biology that were previously intangible because of the lack of sufficiently sensitive analytical methods. Many applications of PCR, including diagnosis of heritable disorders, screening for susceptibility to disease, and identification of bacterial and viral pathogens, require the determination of the nucleotide sequence of amplified DNA fragments. The chapter also discusses double-stranded DNA templates for the sequencing of in vitro amplified DNA. Series: Methods in Enzymology (Book 218) Hardcover: 806 pages Publisher: Academic Press Language: English ISBN-10: 0121821196 ISBN-13: 978-0121821197 Product Dimensions: 1.5 x 6.2 x 9.5 inches Link Download http://nitroflare.com/view/87B4F5AE49A1EB5https://drive.google.com/drive/folders/1yLBzZ1rSQoNjmWeJTZ3WGQHg04L1
