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Gene Transfer and Expression de Michael Kriegler

Gene Transfer and Expression

Autor Michael Kriegler
en Limba Engleză Paperback – 1990
A practical manual of protocols for achieving expression of foreign genes in mammalian cells. It includes some very new techniques such as PCR-based expression. The author gives a theoretical introduction to the protocols and compares the strengths and weaknesses.
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Specificații

ISBN-13: 9781349118939
ISBN-10: 1349118931
Pagini: 260
Ilustrații: XIII, 242 p.
Dimensiuni: 210 x 279 x 14 mm
Greutate: 0.59 kg
Ediția:1990 edition
Editura: Springer Nature B.V.
Locul publicării:London, United Kingdom

Cuprins

Part 1 Overview: general comparison of vectors and DNA transfer procedures; transient and stable expression systems; viral compared to DNA mediated transfer; stratefic overview - matching experimental procedures with goals and probability of success. Part 2 Vectors (DNA): vector components - organization of the most efficient vectors such as pMT2, pCD, SRa, EBV based, BPV based and T7 based vectors; control elements; retroviral vectors and recombinant retroviruses; PCR based expression; homologous recombination and gene replacement. Part 3 Cell lines and maintenance: general tissue culture techniques; growth conditions of most commonly used mammalian cells - COS, 3t3, CV1, CHO, C127, BHK, mouse L cells, Att20, MDCK, lymphoid cells; embryonal stem cells. Part 4 DNA transfer: calcium phosphate; DEAE dextran; protoplast fusion; electroporation; chromosome mediated transfer - DEPC/PEF fusion, gamma irradiation/PEG fusion; selecting transfer technique for cell type and stability of expression. Part 5 Selection and amplification methods: DHER; Ada; Neo; GPT; Hygro; ODC, Asn synthtase, mdr; negative selection protocols (gancyclovir). Part 6 Expression cloning: library construction; transfaction techniques; SIB selection. Part 7 Analysis of nucleic acid: isolation of RNA and DNA; analysis of RNA (traditional and PCR based); analysis of DNA (traditional and PCR based). Part 8 Analysis of protein: radiolabeling, immunoprecipitation and PAGE systems; western immunoblotting; analysis of glycosylation, sulfation and acylation; study of protein processing and secretion. Appendix: reagents - where to obtain and how to prepare; troubleshooting.

Descriere

Descriere de la o altă ediție sau format:
This book is a state-of-the-art catalogue of viral and cis-acting elements and the latest expression vectors for obtaining optimum expression of foreign genes introduced into mammalian cells. It describes in detail the essential techniques for propagating cells and cell lines, DNA transfer, drug selection and gene amplification, expression cloning, subtractive hybridizaton, retro-virus mediated gene transfer.