Protein Expression and Purification Series - Bio-Rad
Protein Expression and Purification Series - Bio-Rad
Protein Expression and Purification Series - Bio-Rad
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<strong>Protein</strong> <strong>Expression</strong> <strong>and</strong> <strong>Purification</strong> <strong>Series</strong><br />
Table of Contents<br />
Table of Contents<br />
Kit Summary<br />
7<br />
Storage Instructions<br />
7<br />
<strong>Series</strong> Inventory Checklist<br />
9<br />
Checklists for Individual Modules<br />
10<br />
Materials Required But Not Supplied<br />
12<br />
Course Objectives<br />
15<br />
Note to the Instructor<br />
16<br />
Timeline for the Laboratory Course<br />
18<br />
Chapter 1: Recombinant <strong>Protein</strong> <strong>Expression</strong> <strong>and</strong> <strong>Purification</strong><br />
33<br />
Chapter 2: Focus Questions<br />
49<br />
Chapter 3A: Advanced Preparation for Centrifugation <strong>Purification</strong> Protocols<br />
57<br />
Cell Culture<br />
57<br />
Cell Lysis<br />
62<br />
<strong>Protein</strong> <strong>Purification</strong> by Centrifugation<br />
66<br />
<strong>Protein</strong> Quantitation<br />
67<br />
SDS-PAGE Electrophoresis to Check <strong>Expression</strong> <strong>and</strong> Purity<br />
68<br />
DHFR Enzyme Activity<br />
71<br />
Chapter 3B: Advance Preparation for Chromatography Instrumentation Protocols 72<br />
Cell Culture<br />
72<br />
Cell Lysis<br />
78<br />
SDS-PAGE Electrophoresis to Check <strong>Expression</strong> <strong>and</strong> Purity<br />
81<br />
<strong>Protein</strong> <strong>Purification</strong> with Chromatograpy Instrumentation<br />
84<br />
DHFR Enzyme Activity<br />
89<br />
Chapter 4: Culturing, <strong>Expression</strong>, Lysis <strong>and</strong> SDS-PAGE Analysis for 11 ml Cultures<br />
<strong>and</strong> <strong>Purification</strong> Student Protocol for Centrifugation Process<br />
91<br />
Cell Culture, Subculture <strong>and</strong> Induction<br />
91<br />
Lysing <strong>and</strong> Pelleting Cells<br />
97<br />
Preparing SDS-PAGE Samples <strong>and</strong> SDS-PAGE Electrophoresis<br />
99<br />
Spin Chromatography <strong>Purification</strong><br />
101<br />
<strong>Protein</strong> Quantitation<br />
104<br />
SDS-PAGE Electrophoresis to Check <strong>Expression</strong> <strong>and</strong> Purity<br />
106<br />
Workflow<br />
108<br />
Chapter 5: Culturing, <strong>Expression</strong>, Lysis <strong>and</strong> SDS-PAGE Analysis for 100 ml Cultures<br />
for Chromatography Instrumentation <strong>Purification</strong><br />
118<br />
Cell Culture, Subculture <strong>and</strong> Induction<br />
118<br />
Lysing <strong>and</strong> Pelleting Cells<br />
124<br />
Preparing SDS-PAGE Samples <strong>and</strong> SDS-PAGE Electrophoresis<br />
126<br />
Workflow<br />
130<br />
Chapter 6: H<strong>and</strong>packing a Glass Econo-Column Column attached to a Flow Adaptor 134<br />
Chapter 7: <strong>Purification</strong> Protocol for <strong>Bio</strong>Logic LP System<br />
139<br />
Preparing the <strong>Bio</strong>Logic LP System for Use<br />
139<br />
Installing the Prepacked Column—<strong>Bio</strong>-Scale Mini Profinity IMAC Cartridge 142<br />
Start Method<br />
149<br />
Cleaning the <strong>Bio</strong>Logic LP System After Use<br />
151<br />
Preparing for SDS-PAGE Analysis<br />
152<br />
5<br />
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