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Generation of cDNA Libraries: Methods and Protocols: Methods in Molecular Biology, cartea 221

Editat de Shao-Yao Ying
en Limba Engleză Paperback – 10 noi 2010
Since its invention and subsequent development nearly 20 years ago, po- merase chain reaction (PCR) has been extensively utilized to identify numerous gene probes in vitro and in vivo. However, attempts to generate complete and full-length complementary cDNA libraries were, for the most part, fruitless and remained elusive until the last decade, when simple and rapid methods were developed. With current decoding and potential application of human genome information to genechips, there are urgent needs for identification of functional significance of these decoded gene sequences. Inherent in bringing these app- cations to fruition is the need to generate a complete and full-length cDNA library for potential functional assays of specific gene sequences. Generation of cDNA Libraries: Methods and Protocols serves as a laboratory manual on the evolution of generation of cDNA libraries, covering both ba- ground information and step-by-step practical laboratory recipes for which p- tocols, reagents, operational tips, instrumentation, and other requirements are detailed. The first chapter of the book is an overview of the basics of generating cDNA libraries, which include the following: (a) the definition of a cDNA library, (b) different kinds of cDNA libraries, (c) differences between methods for cDNA library generation using conventional approaches and novel stra- gies, including reverse generation of RNA repertoires from cDNA libraries, and (d) the quality of cDNA libraries.
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Specificații

ISBN-13: 9781617373336
ISBN-10: 1617373338
Pagini: 338
Ilustrații: XIV, 338 p.
Dimensiuni: 178 x 254 x 19 mm
Greutate: 0.61 kg
Ediția:Softcover reprint of hardcover 1st ed. 2003
Editura: Humana Press Inc.
Colecția Humana
Seria Methods in Molecular Biology

Locul publicării:Totowa, NJ, United States

Public țintă

Research

Cuprins

Complementary DNA Libraries.- Rapid Amplification of cDNA Ends.- cDNA Generation on Paramagnetic Beads.- Construction of a Normalized cDNA Library by mRN-cDNA Hybridization and Subtraction.- Amplification of cDNA Ends Using PCR Suppression Effect and Step-Out PCR.- Use of Inverse PCR to Clone cDNA Ends.- Construction of Size-Fractionated cDNA Library Assisted by an In Vitro Recombination Reaction.- Construction of a Full-Length Enriched and a 5?-End Enriched cDNA Library Using the Oligo-Capping Method.- cDNA Library Construction Using In Vitro Transcriptional Amplification.- Amplification of Representative cDNA Pools from Microscopic Amounts of Animal Tissue.- Single-Cell cDNA Library Construction Using Cycling aRNA Amplification.- mRNA/cDNA Library Construction Using RNA—Polymerase Cycling Reaction.- Quality Assessment of cDNA Libraries.- Assessment of the Quality of mRNA Libraries by Agarose Gel Electrophoresis.- PACS RT-PCR.- Single-Cell mRNA Library Analysis by Northern Blot Hybridization.- Generation of cDNA Libraries for Profiling Gene Expression of Given Tissues or Cells.- Screening Poly [dA/dT(?)] cDNA for Gene Identification.- Generation of Longer cDNA Fragments from SAGE Tags for Gene Identification.- Generation of Full-Length cDNA Libraries Enriched for Differentially Expressed Genes.- Subtractive Hybridization for the Identification of Differentially Expressed Genes Using Uraci-DNA Glycosylase and Mung-Bean Nuclease.- Subtractive Cloning of Differential Genes Using RNA-PCR.- Strategy for Construction of a cDNA Encoding a Repetitive Amino Acid Sequence.- Preparing Lambda Libraries for Expression of Proteins in Prokaryotes or Eukaryotes.- Peptide Library Construction from RNA-PCR-Derived RNAs.- Identifying Interacting Proteins in an Escherichiacoli-Based Two-Hybrid System.- Future Perspectives.

Recenzii

...the ready-for-use protocols are excellent...A good buy. - Cell Biochem Funct

Textul de pe ultima copertă

With the decoding of the human genome and the potential application of its information to gene chips, the ability to generate cDNA libraries for functional assays of specific gene sequences has become critical in gene manipulation and genetic research. In Generation of cDNA Libraries: Methods and Protocols, expert researchers and inventors in the field describe their own proven techniques for generating cDNA/mRNA libraries to identify the functions of specific decoded gene sequences. A wide variety of techniques is presented for enhancing the generation of complete and full-length libraries, and for confirming the quality of the cDNAs generated. Among the applications detailed are electrophoresis, Northern blotting, single-cell microarray analysis, subtractive hybridization, subtractive cloning, gene cloning, and peptide library generation. Each method includes background information, step-by-step instructions, a list of reagents, operational tips, and notes on instrumentation. A background tutorial provides essential information: the definition of a cDNA library, the various types of cDNA libraries, the differences between methods for cDNA library generation using either conventional approaches or novel strategies, and the quality of cDNA libraries.
State-of-the-art and highly practical, Generation of cDNA Libraries: Methods and Protocols provides a variety of powerful and readily reproducible techniques for the generation of the entire range of complete, full-length cDNA/mRNA libraries needed in today's forefront genetic research.

Caracteristici

Includes supplementary material: sn.pub/extras