Methods in Molecular Biology and Protein Chemistry: Cloning and Characterization of an Enterotoxin Subunit

Methods in Molecular Biology is a discovery–based guide that will enable students to master current molecular biology and biochemistry techniques as an integrated part of a research project. Written in an accessible style, this book adopts a practical hands–on approach to encourage the reader to explore and experiment independently. The book is designed to support a one semester laboratory course in which students participate in an ongoing research project, give a research presentation on a topic related to the project, and write a research article suitable for publication.
Methods in Molecular Biology features:
∗ a carefully structured sequence of experiments that enables students to develop key laboratory skills whilst contributing towards a live, research project.
∗ a mid–term mock exercise that encourages students to test test their undertanding half way through the book, assess what they have learnt, utilize the primary literature and propose additional research applications and methods.
∗ a supplementary Website, that includes:
guidelines on searching the primary literature using internet and library search tools
corrections,additions, and troubleshooting suggestions
advice on; how to prepare laboratory notebooks, present oral reports, record keeping and report writing
suggestions for future projects based on modified text protocols
This book will be invaluable for undergraduate students of biology, biochemistry microbiology and biomedical sciences. It will also be useful for those students taking chemistry, chemical engineering and animal and plant science courses looking to improve their practical experience in the lab.

Spis treści:
Preface
Introduction and Background
Part 1 Molecular Biology
Experiment 1 Instructions for Pipetman Use
Experiment 2 Preparation of Plasmid Template DNA
Experiment 3 Estimation of Plasmid DNA Concentration
PCR Amplification of LTB Gene from Plasmid DNA
Experiment 4 Agarose Gel Eletrophoresis
Recovery of PCR Product
Experiment 5 Restriction Digest of LTB Insert
Plate Preparation
Experiment 6 Purification of Digested Insert and Vector by Agarose Gel Eletrophoresis
Recovery of Digested Insert and Vector
Experiment 7 Determination of DNA Concentrations
Ligation of Insert into Vector
Experiment 8 Transformation of Host Cells by Construct
Experiment 9 Colony–pick PCR to Check for Presence of Insert
Streak Plates with Potential Constructs
Inoculate Cultures/Run PCR
Experiment 10 Agarose Gel Electrophoresis to Verify Presence of Insert
Purification of the Plasmid Construct using a Plasmid Miniprep Kit
Experiment 11 DNA Concentration Determination
Ethanol Precipitation of Plasmid DNA
Sequencing Reaction
Experiment 12 Purification of Extension Reaction Products
Sequencing Gel Demonstration
Experiment 13 Analysis of Sequencing Data
Verification of Insert Sequence
Experiment 14 Gene Expression
Transformation into Expression Host
Experiment 15 Induction of LTB Gene Expression
Part 2 Protein Chemistry
Experiment 16 SDS–PAGE of Induction Time Course
Transfer of Protein to Membrane for Western Blot
Experiment 17 Visualization of Western Blot
Experiment 18 Large Scale Culture Preparation
Experiment 19 Isolation of LTB by Affinity Chromatography
Experiment 20 SDS–PAGE of Column Fractions
Immunoblot to verify Presence of LTB
Experiment 21 Protein Concentration
Protein Crystallization
Experiment 22 Setting up an ELISA Microtiter Plate
Experiment 23 ELISA of a Panel of Ganglioside Ligands
Experiment 24 Protein Structure Determination: X–ray Diffraction Techniques
Experiment 25 Characterization of a Protein Crystal
Experiment 26 Primer Design

Okładka tylna:
Methods in Molecular Biology is a discovery–based guide that will enable students to ma ster current molecular biology and biochemistry techniques as an integrated part of a research project. Written in an accessible style, this book adopts a practical hands–on approach to encourage the reader to explore and experiment independently. The book is designed to support a one semester laboratory course in which students participate in an ongoing research project, give a research presentation on a topic related to the project, and write a research article suitable for publication.
Methods in Molecular Biology features:
∗ a carefully structured sequence of experiments that enables students to develop key laboratory skills whilst contributing towards a live, research project.
∗ a mid–term mock exercise that encourages students to test test their undertanding half way through the book, assess what they have learnt, utilize the primary literature and propose additional research applications and methods.
∗ a supplementary Website, that includes:
guidelines on searching the primary literature using internet and library search tools
corrections,additions, and troubleshooting suggestions
advice on; how to prepare laboratory notebooks, present oral reports, record keeping and report writing
suggestions for future projects based on modified text protocols
This book will be invaluable for undergraduate students of biology, biochemistry microbiology and biomedical sciences. It will also be useful for those students taking chemistry, chemical engineering and animal and plant science courses looking to improve their practical experience in the lab.