Transgenic Core

Director: Arlene Sharpe


 
 
Core Summary:

The BWH Transgenic Core, established in 1992 by Dr. Arlene Sharpe, provides gene targeting and microinjection services using state-of-the-art facilities and equipment.

The Core has extensive experience generating transgenic mice using DNA constructs (including BAC DNAs) and lentiviral constructs. We also have a track record of success generating KO/KI mice by performing gene targeting, or by expanding targeted ES Cells from consortia.

The Core works with investigators to achieve optimal outcomes for each project.

Personnel/Contact Information:

Director: Arlene Sharpe, M.D, Ph.D.
Phone: (617) 432-6569
Fax: (617) 432-6570
Email:
arlene_sharpe@hms.harvard.edu

Supporting Staff:

Transgenic Specialist: Lina Du, M.D.
Phone: (617) 432-6227
Email: lina_du@hms.harvard.edu

ES Cell Culture Technician: Jennifer Lesko
Phone: (617) 432-6182
Email: bwhtransgeniccore@gmail.com

Services:

    Click for New Services Flyer

  • Microinjection of DNA constructs (including BACs) into pronuclei for generating transgenic mice (FVB or C57 BL/6 strain background)
  • Microinjection of embryonic stem (ES) cells into blastocysts for generating knockout mice
  • ES cell transfection for generating ES cell clones with the targeted combination event
  • Lentiviral transgenics
  • Microinjection of IPS

 

FY14 User Fees (effective 02/19/2014):

Service

(See Applications in Outline & Timeline Section)
Internal Users
External Users
Partners
Non-Partners AMC & Non-Profit
For-Profit & Industry
Lentiviral Injection
$3,938
$5,671
$6,262
DNA (FVB) Pronuclear Injection
$4,312
$6,210
$6,857
DNA (C57) Pronuclear Injection
$4,100
$5,904
$6,519
DNA Injection & Microdissection
$2,077
$2,991
$3,303
Additional DNA Injection
$2,261
$3,256
$3,595
BAC Injection
$4,700
$6,768
$7,473
ES Cell Injection (129)
$2,269
$3,268
$3,608
ES Cell Injection (B6)
$2,345
$3,377
$3,729
Tail Biopsy
$175
$252
$279
Electroporation (129) ES
$2,877
$4,143
$4,575
Electroporation (B6) ES
$3,223
$4,642
$5,125
ES Expansion
$680
$980
$1,082
ES Cell Prep for Injection
$263
$379
$419
Balb C
$5,325
$7,668
$8,467
IPS
$2,935
$4,227
$4,667
CRISPR Injection
$2,345
$3,377
$3,729
Sperm Cryopreservation
$400
$576
$636
Embryo Cryopreservation
$550
$792
$875
Thawing and Implantation of Embryos
$550
$792
$875
Fresh Sperm IVF
$1,400
$2,016
$2,226
Frozen Sperm IVF
$1,400
$2,016
$2,226
Exportation Fee
TBD
TBD
TBD
Animal Per Diem
TBD
TBD
TBD


Outline & Timeline of Project:

Our ES facility will generate targeted ES cell lines with KO/KI constructs provided by the investigator. Upon receipt of the construct and application form, an electroporation date will be scheduled. The Core Director, Dr. Arlene Sharpe, will prioritize the service requests according to the difficulty of the project and workload at the time of receipt.

  1. Transfection of your purified linearized DNA construct into ES cells, JM8 (C57 BL/6), J1 (129/SV) and Bruce 4 (C57 BL/6J).
  2. After the electroporation and selection, clones will be picked (192), cultured in multi-well plates and replicated. The clones will then be frozen in order to expand the positive clones after screening. This part of the process usually takes 3 weeks to complete.
  3. The investigator will be notified and given DNA isolated from each clone suitable for screening by Southern and PCR.
  4. Since storage of frozen cells is not optimal, it is strongly suggested that investigators return the screening results to the core within 3 months of receipt of the DNA samples.
  5. After receiving screening results, up to 12 ES positive clones will be expanded, split for DNA, and frozen down. The investigator will then receive DNA from each clone for reconfirmation of targeted recombination. This process usually takes 2 weeks.
  6. After reconfirmation, the best clone will then be scheduled for injection into blastocysts. Chimeric mice will be given to the investigator.

 

Please complete the appropriate application forms below for your service request:

CRISPR Application

Cryopreservation Application

Fresh Frozen Sperm IVF Application

Thawing and Implantation of Embryos Application

Gene Targeting Application

Injection of DNA into Pronuclei

ES Cells into Blastocysts

Completed applications can be sent to bwhtransgeniccore@gmail.com or dropped off to Jennifer Lesko at NRB Lab #837 before your project is started.

 


 

 
 
     
 

 

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