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Custom CRISPR Validation Service

Performed a CRISPR experiment, but unsure whether you have the right cell clone for your research? Looking for a third party to confirm your CRISPR results for publication? abm is proud to offer a variety of CRISPR validation options – from sgRNA efficiency screening, genomic cleavage assay, analysis by Sanger sequencing or NGS, to protein-level validation – to provide thorough validation of your CRISPR results.

 

 

“We are quite happy with the knockout cells that you made. This was a tricky knockout and hemizygous removal was much more prevalent than the double knockout, but you did it. In addition to your excellent genomic sequence characterization of both mutant LIF alleles, I have confirmed in our lab that the tumor cells have lost all mouse LIF production.””

Dr. Robert Jackman, Boston University, Mouse LIF CRISPR Knock Out C26 Tumor Cell Line Generation and Screening Service

Additional Resources:

 

Service Details

Core Services

SERVICE NAME UNIT CAT. NO.

sgRNA Efficiency Screening Service

List price includes screening up to a set of 4 sgRNAs with 1 set of primers.

Additional services:

  • $ 142.50 per additional sgRNA tested (C531)
  • $ 148.50 per additional primer design & synthesis required (C532)
 1 Service C526

Mismatch Cleavage Detection Service (Surveyor/T7E1 Assay)

*Minimum sample requirement: 8 samples

 1 Sample C527

SpeedySeq Sequencing of Target Region

PCR amplicon of target region is subcloned into a vector and 10 isolated colonies are sequenced to investigate the exact edited sequence.


*Sample is based on sequencing one target region, for sequencing multiple regions in the same sample, additional charges will apply.

 1 Sample C528

SpeedySeq Sequencing of PCR Amplicon

PCR amplicon of target region is directly sequenced without subcloning to access whether edits are present.

*Sample is based on sequencing one target region, for sequencing multiple regions in the same sample, additional charges will apply.

 1 Sample C529
Validation Service by Western Blot (Up to 10 Clones) 1 Service C145
CRISPR-Cas9 Amplicon-Seq 1 Sample IA00100

Add-On Services

SERVICE NAME UNIT CAT. NO.
gDNA Isolation from cell pellet Per Cell Pellet C530

 

Additional Info

Service Selection Guide

 
SERVICE  PURPOSE MATERIALS TO SUBMIT DELIVERABLES
sgRNA Efficiency Screening Service Validate sgRNA designs in vitro prior to performing CRISPR experiments

Cell pellet or isolated gDNA from target cells. Alternatively, abm can provide 293T gDNA for free

sgRNA sequence information

 Report with Cleavage assay gel image showing the cleavage efficiency of the different sgRNA designs
Mismatch Cleavage Detection Service (Surveyor/T7E1 Assay) Upon infection, determine if the polyclonal pool of cells contains edits

gDNA or cell pellet of CRISPR-edited samples

gDNA or cell pellet from Wildtype cell sample

 Cleavage detection assay gel image
SpeedySeq Sequencing of Target Region Provides detailed sequence information of the CRISPR target region

gDNA or cell pellet from CRISPR-edited cells

Reference sequence

 10 sequencing results/sample, service report showing alignment to reference sequence
SpeedySeq Sequencing of PCR Amplicon Used for preliminary screening to determine whether edits are present. Clean sequencing results can be seen for monoclonal samples. If messy sequencing results are seen at the expected sgRNA cleavage point, then the sample likely contains a mixture of edits/ sequences.

gDNA or cell pellet from CRISPR-edited cells

Reference sequence

 1 sequencing result/sample, service report showing alignment to reference sequence
Validation Service by Western Blot (Up to 10 Clones) Protein-level validation

Protein lysate from wild-type cells (non-edited sample)

Protein lysate from edited cells

WB positive control

100 μg of pre-validated antibody

 Custom report on Western blot results
CRISPR-Cas9 Amplicon-Seq Can validate multiple samples at once and provides deeper sequencing coverage by NGS

2 ug gDNA/sample

 FASTQ data
Off-Target Analysis by Whole Genome Sequencing Determines off-targets

Customer must submit a minimum of 2 samples, 1 Wildtype for control 1 edited sample, 2 ug gDNA/sample

 FASTQ data

 

sgRNA Efficiency Screening Service:

  1. Primer design and synthesis (1 set of primers)
  2. sgRNA template synthesis
  3. In vitro transcription of sgRNAs
  4. Amplification of DNA target(s) from gDNA
  5. In vitro cleavage assay with Cas Nuclease Protein (please choose one from the available list: saCas9, spCas9, fnCpf1, asCpf1)

Deliverables: Report with cleavage assay gel image
Lead time: 2-4 weeks

Mismatch Cleavage Detection Service (Surveyor/T7E1 Assay):

  1. Primer design and synthesis (up to 2 sets of primers tested)
  2. Amplify region of interest from gDNA
  3. Mismatch cleavage detection assay, 1 reaction/sample

Deliverables: Report with cleavage detection assay gel image
Lead time: 3-5 weeks

SpeedySeq Sequencing of Target Region

  1. Primer design and synthesis (up to 2 sets of primers tested)
  2. Amplification of DNA target from gDNA
  3. Subcloning
  4. Culture and isolate 10 clones per sample
  5. Sanger sequencing of clones (target region sequenced is up to 700bp)
  6. Alignment to reference sequence

Deliverables: 10 sequencing results/sample, service report showing alignment to reference sequence
Lead time: 3-5 weeks

SpeedySeq Sequencing of PCR Amplicon

  1. Primer design and synthesis (up to 2 sets of primers tested)
  2. Amplification of DNA target from gDNA
  3. Sanger sequencing of PCR product
  4. Alignment to reference sequence

* Target region sequenced is up to 700bp
Deliverables: 1 sequencing result/sample, service report showing chromatogram and alignment to reference sequence
Lead time: 2-4 weeks

Protein CRISPR Validation Service

  1. Western blot will be performed on wild-type sample, CRISPR-edited samples, and the positive control using pre-validated antibody.

Deliverables: Custom report on Western blot results
Lead time: 2-4 weeks
*All lead times specified are estimates and based on receipt of gDNA or protein lysate samples. If a gDNA isolation service is required, 1 additional week will be added.

Sample Submission Guideline

SAMPLE FORMAT DETAILS
gDNA
  • Isolated gDNA from at least 1 million cells/sample
  • The gDNA template must be non-degraded and unamplified. If samples are found to be degraded, the customer will be asked to resend the gDNA.
  • Avoid repeated freeze thawing and store in nuclease-free centrifuge tubes in Tris-HCl buffer (pH 8.0) at -20°C.
  • Ship samples on blue ice packs to keep samples cold during transport
Cell pellet
  • Cell pellet containing at least 1 million cells/sample*
  • Ship samples in dry ice to maintain sample quality during transit* gDNA Isolation service charges will apply

 

 

Disclaimer and Policies

abm is not responsible for storage of any reagents associated with the customer (cell line, DNA, PCR product and cloned plasmids) after one month of the report delivery.

Our goal is always to deliver high-quality validation data to aid you in your research and downstream application. However, CRISPR validation service results are dependent on the sample quality submitted by customers and as such, abm is not in a position to guarantee the final results obtained and full service charges will apply for the services performed. By placing an order with abm, customer agrees to this term.

All services are for research use only.

 

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