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Cellartis iPSC rCas9
Takara Bio Cellartis iPSC rCas9 Manuals
Manuals and User Guides for Takara Bio Cellartis iPSC rCas9. We have
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Takara Bio Cellartis iPSC rCas9 manual available for free PDF download: User Manual
Takara Bio Cellartis iPSC rCas9 User Manual (24 pages)
Electroporation and Single-Cell Cloning System
Brand:
Takara Bio
| Category:
Science Education products
| Size: 0.98 MB
Table of Contents
Table of Contents
2
Introduction
4
Figure 1. Schematic of Crispr/Cas9-Based Editing to Create Gene Knockouts and Knockins
5
II. List of Components
6
List of Components
6
Additional Materials Not Supplied
7
Required
7
Recommended
7
IV. General Considerations
8
Cloning System
8
General Considerations
8
Storage and Handling
8
Transferring Human Ips Cells to the DEF-CS Culture System
8
Complete Experimental Workflow
9
In Vitro Transcription of an Sgrna Containing the Desired Target Sequence
9
Protocol Overview
9
Generating the DNA Template
10
Figure 5. Designing a Forward PCR Primer to Generate a DNA Template for an Sgrna Containing Your Target Sequence
11
Performing the in Vitro Transcription (IVT) Reaction
12
Figure 7. Sgrna Yield over Time
13
Purifying the Transcribed Sgrna
14
Table I. Examples of Sgrna Concentration and Yield Using Different Elution Volumes
14
Editing Hips Cells by Electroporation
15
Protocol Overview
15
Preparing Media and Cells
15
Table II. Preparation of Coating Solution for a 48-Well Plate
16
Table III. Preparation of DEF-CS Medium for Dissociation and Recovery
16
Preparing the Rcas9/Sgrna RNP Complex
17
Electroporating Cells Using the Neon Transfection System
18
Table IV. Workflow for Single-Cell Cloning and Expansion Using the Cellartis Ipsc Rcas9 Electroporation and Single-Cell
19
Single-Cell Seeding into a 96-Well Plate
20
Single-Cell Cloning of Edited Cells
19
Protocol Overview
19
Table V. Preparation of Coating Solution for a 96-Well Plate
20
Table VI. Preparation of Medium for Dissociation and Single-Cell Seeding
21
Culturing Single-Cell Colonies
22
Table VII. Preparation of Coating Solution for a 48-Well Plate
23
Passaging Cells from the 96-Well Plate to a 48-Well Plate
23
Coating a 48-Well Plate
23
Preparing DEF-CS SCC Medium for Passaging
23
Passaging
23
Scaling up from the 48-Well Plate
24
References
24
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