Generation of cDNA Libraries

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.

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.