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Guide-it Indel Identification Kit

The Guide-it Indel Identification Kit provides a complete workflow for identifying the variety of insertions and deletions (indels) created in a cell population using nuclease-based genome editing technologies, including CRISPR/Cas9, TALENs, and zinc finger nucleases. The kit contains all of the components needed to amplify, clone, and prepare modified target sites for DNA sequence analysis. The indel identification protocol in this kit has been optimized to provide quick and reliable results. First, Terra PCR Direct polymerase is used to amplify a genomic DNA fragment containing the target site directly from crude cell lysates. The resulting amplified fragments are cloned directly, without restriction digestion, into a pre-linearized pUC19 vector using the In-Fusion Cloning system. After transformation into Stellar Competent Cells, colony PCR is used to amplify the target region from the plasmid. DNA sequence analysis of the PCR products is then used to identify indels generated at the targeted genomic site.

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Cat. #	Product	Size	Price	License	Quantity	Details
631444	Guide-it™ Indel Identification Kit	10 Rxns	USD $485.00
The Guide-it Indel Identification Kit is used for characterization of insertions and deletions (indels) generated by gene editing tools, such as CRISPR/Cas9. This kit contains all of the components needed to amplify, clone, and prepare modified target sites for DNA sequence analysis. This kit uses Terra PCR Direct to amplify targets directly from crude genomic DNA extracts. The resulting pool of fragments, which may contain a variety of indels, are cloned into a prelinearized pUC19 vector using the In-Fusion cloning system. Colony PCR of individual clones using Terra PCR Direct followed by DNA sequencing allows indel characterization. Notice to purchaser Our products are to be used for Research Use Only. They may not be used for any other purpose, including, but not limited to, use in humans, therapeutic or diagnostic use, or commercial use of any kind. Our products may not be transferred to third parties, resold, modified for resale, or used to manufacture commercial products or to provide a service to third parties without our prior written approval. Documents Components Image Data Resources Back Identification of insertions and deletions (indels) in the CD81 gene after CRISPR/Cas9 targeting Identification of insertions and deletions (indels) in the CD81 gene after CRISPR/Cas9 targeting. HeLa cells were transfected with plasmids encoding Cas9 and an sgRNA targeting the CD81 gene. The Guide-it Indel Identification Kit was used to prepare CD81 target sites for DNA sequence analysis. The sequencing data from six clones was aligned with the wild-type sequence, revealing a broad range of indels in the CD81 gene. Back The Guide-it Indel Identification Kit provides a complete workflow for identifying the variety of insertions and deletions (indels) introduced by nuclease-based genome editing The Guide-it Indel Identification Kit provides a complete workflow for identifying the variety of insertions and deletions (indels) introduced by nuclease-based genome editing. The protocol uses direct PCR to amplify a genomic DNA fragment (~500 to 700 bp) containing the target site directly from crude cell lysates (step 1). The resulting amplified fragments contain a pool of edited target sites from individual cells. These PCR products are cloned directly into a pre-linearized pUC19 vector using the In-Fusion Cloning system (step 2). After transformation of an optimized E. coli strain, colony PCR is used to amplify the target site from the plasmid (step 3). DNA sequencing is then used to identify the different indels generated at the targeted genomic site (step 4) Back The CRISPR/Cas9 system, a simple, RNA-programmable method to mediate genome editing in mammalian cells The CRISPR/Cas9 system, a simple, RNA-programmable method to mediate genome editing in mammalian cells. The CRISPR/Cas9 system relies on a single guide RNA (sgRNA) directing the Cas9 endonuclease to induce a double strand break at a specific target sequence three base-pairs upstream of a PAM sequence in genomic DNA. This DNA cleavage can be repaired in one of two ways: 1) nonhomologous end joining, (NHEJ) resulting in gene knockout due to error-prone repair (orange), or 2) homology-directed repair (HDR), resulting in gene knockin due to the presence of a homologous repair template (purple). Back 631444: Guide-it Indel Identification Kit

Overview

Streamlined method for characterizing the variety of indels introduced by gene editing technologies
Ultrafast protocol includes Terra PCR Direct Polymerase for PCR amplification directly from cells and the In-Fusion Cloning system for ligation-free cloning in 15 minutes
Ideal for identifying indels caused by Cas9/CRISPR, TALENs, or zinc-finger nuclease-based gene editing in a cell population
Complete kit contains all of the components needed to amplify, clone, and prepare modified target sites for DNA sequence analysis

More Information

Applications

Detecting insertions and deletions introduced in mammalian genomic DNA
Characterizing the variety of indels that are present in a cell population after nuclease-based gene editing

Components

Guide-it Indel Identification Components
- 110 µl Terra PCR Direct Polymerase Mix
- 3 x 1 ml 2X Terra PCR Direct Buffer (with Mg²⁺, dNTP)
- 400 µl Extraction Buffer 1
- 40 µl Extraction Buffer 2
- 10 µl pUC19 Cloning Vector, linearized (50 ng/µl)
- 200 µl Colony PCR Forward Primer (15 µM)
- 200 µl Colony PCR Reverse Primer (15 µM)
- 6 x 1 ml PCR-Grade Water

In-Fusion HD Cloning Kit
NucleoSpin Gel and PCR Clean-up kit
Stellar Competent Cells

Additional product information

Please see the product's Certificate of Analysis for information about storage conditions, product components, and technical specifications. Please see the Kit Components List to determine kit components. Certificates of Analysis and Kit Components Lists are located under the Documents tab.

CRISPR/Cas9 information

Choosing sgRNA design tools

Browse a collection of sgRNA design tools for Cas9-based genome editing experiments.

Choosing a target sequence for CRISPR/Cas9 gene editing

Learn how to design sgRNA sequences for successful gene editing.

The CRISPR/Cas9 system for targeted genome editing

Overview of CRISPR/Cas9 system for genome editing.

CRISPR/Cas9 genome editing tools

An overview of tools available for each step in a successful genome editing workflow.

Gene editing technical notes

Delivery of Cas9 and sgRNA to mammalian cells using a variety of innovative tools.

SNP engineering application note

Learn about a simple assay for sensitive detection of single-nucleotide substitutions in bulk-edited or clonal cell populations.

CRISPR/Cas9 gesicles overview

Learn about Guide-it CRISPR/Cas9 Gesicle Production System components and workflow.

CRISPR library screening webinar

Watch this webinar to learn how you can perform genome-wide lentiviral sgRNA screens easily.

Choosing an HDR template format

Watch a webinar on how to choose the right HDR template for knockin experiments.

Guide-it SNP Screening Kit FAQs

Get answers to frequently asked questions and view a video explaining the enzymatic assay.

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Guide-it Indel Identification Kit

Notice to purchaser

Identification of insertions and deletions (indels) in the CD81 gene after CRISPR/Cas9 targeting

The Guide-it Indel Identification Kit provides a complete workflow for identifying the variety of insertions and deletions (indels) introduced by nuclease-based genome editing

The CRISPR/Cas9 system, a simple, RNA-programmable method to mediate genome editing in mammalian cells