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Fees - Duke Cancer Institute Shared Resource

Below is a general description of our services and approved SOM FY 2019 fees.  Please contact Cheryl Bock for ES Cell and Embryo Services or Gary Kucera for Molecular Biology Services.  To download a request form click here.

 

ES Cell/Gene-Targeting Services

Generate HITI-Based Targeting Vector: $3,780

 Includes: Sequence verified, linearized, targeting vector DNA (Knockout, knockin, cKO, cKI model;) plus validated PCR assay and MOCK control DNA for screening  targeted ES cell clones.

 ES Cell Transfection of targeting vector: $3,800

 We have the best results with G4 ES cells which are a 129/B6N hybrid ES line we have obtained by MTA from Mount Sinai Hospital in Toronto.  Homozygous ES cells that are: 129S6/SvEvTac & also B6N ES cells can be used for transfection but this must be discussed before the targeting construct is made.

ES Cell Screening of Putative Targeted Clones: $1,189

 Up to 18 PCR screened positive ES cell clones will be processed and validated using PCR and sequencing.

 o    DNA Preparation only  from ES clones by RGEST for PI Lab: $500

Morulae Microinjection of Positive ES Cell clones: $4,000

 G4 ES cells of positive clones confirmed by the contracting lab or by the RGEST will be microinjected into ICR morulae to produce chimeric mice. Blastocysts from C57BL/6J will be injected for 129 or B6 ES cell lines. Blastocyst injection is a less efficient method to produce chimeric mice.

  

CRISPR/Cas9 Reagents and In-Vivo Genome Editing Services

Guide RNA design and cloning: $3,290

Design and clone three independent guide RNAs into pX330 (or other) for one target locus.  Sequence Verify.

 

sgRNA Production via In-Vitro Transcription: $1,300

Required for ALL CRISPR electroporation  requests, including Client-provided guide sequences.

 RGEST prepares electroporation ready samples, No Exceptions!

 

Embryo Electroporation of CRISPR Reagents: $3,500

PLEASE NOTE:  All KI models (point mutation, inserted sequence, etc.) require the RGEST to perform Founder Analysis.

Validated sgRNA (RGEST provided reagent only) + repair/donor oligo (if required) + cas9 protein.

 

 Founder Analysis, Initial: $1,000

 Genomic DNA preparation and sequence analysis of targeted locus from up to 24 founder animals to identify Indels. Founder Analysis by RGEST is a REQUIRED step for any KI project.

 

Founder analysis, Advanced, Allelic Subcloning: $1,875.

Subclone and sequence individual targeted alleles from up to 6 founders to identify/verify mutation

  

Pronuclear Injection of Client provided DNA for Transgenic Mouse Production Or BAC Transgene (RGEST or PI Lab)

BAC Transgene Construction: $4,350

 BAC Transgene construction, including removal of vector-specific LoxP site, Maxi prepared and linearized BAC DNA, analysis of final clone via pulse-field gel electrophoresis.

DNA Microinjection to Produce Transgenic Mice: $3,500

One-cell mouse embryos generated from a standard hybrid B6SJLF1/J cross, most efficient.

C57BL/6J embryos can also be used, but efficiency of transgenic production is much less than in hybrid background.  We provide NO Guarantees that founders will be produced.