CRISPR/Cas9 Mouse Targeting Core (UPenn)

 

MISSION STATEMENT: Our main goal has been always to streamline procedures that facilitate investigators' use of the CRISPR/Cas9 genome editing technology for the rapid and economic generation of novel mouse genetic tools. Our core has had a significant positive impact upon the community. Since we established this technology in campus, we have generated over 200 new mouse models for 115 users at UPenn/CHOP, nationwide and abroad. In addition, our R&D efforts in this area should enable UPenn/CHOP to remain at the forefront of this technology.

 

DISCLAIMER: However, we want to stress one more time that the CRISPR/Cas9 Mouse Targeting Core at the University of Pennsylvania provides an academic service to the community, and even though our overall rate of success is high, NEITHER A SUCCESSFUL TARGETING NOR GERM-LINE TRANSMISSION CAN BE GUARANTEED BEFORE HAND. We strongly advise our users to seek the services of a company when a POSITIVE TARGETING MUST BE GUARANTEED BEFORE HAND.

 

ONLY e-mail INQUIRIES, PLEASE! Do not submit your request for CRISPR/Cas9 services before meeting with us!

 

OVERVIEW OF SERVICES - OFFERED CRISPR/Cas9 TARGETING STRATEGIES

(LAST UPDATE ON 05/21/2019)

 

*KNOCK-OUT MICE (KO): SUCCESS RATE = ~85% of the projects with at least one positive (targeted) founder mouse. These data include embryonic-lethal targeting strategies

Generation of knock-out (KO) mice usually requires the design of the targeting strategy, production of Cas9 mRNA/protein and 2 or more gRNAs, microinjection of one-cell embryos, and screening of founder mice. Double, triple or quadruple KO mice of different genes can be accomplished with one injection.

 

*KNOCK-IN MICE - SMALL SEQUENCES (Small-KI): SUCCESS RATE = ~50% of the projects with at least one positive (targeted) founder mouse. These data include embryonic-lethal targeting strategies.

Generation of small knock-in (Small KI) mice containing small sequences (e.g. point mutations, tags, flags, restriction sites, etc.) usually requires the design of the targeting strategy, production of Cas9 mRNA/protein and a gRNA, preparation of donor ssDNA templates, microinjection of one-cell embryos, and screening of founder mice.

 

*CONDITIONAL KNOCK-OUT MICE (cKO): SUCCESS RATE = ~25% of the projects with at least one positive (targeted-CIS) founder mouse.

Generation of conditional knock-out (cKO) mice usually requires the design of the targeting strategy, production of Cas9 mRNA/protein and 2 gRNAs, preparation of 2 donor ssDNA templates containing loxp sites, microinjection of one-cell embryos, and screening of founder mice.

Only ONE (1) loxP site inserted (either 5’ or 3’ loxP site): 45% of the projects with at least one positive (targeted) founder mouse.

Two loxP sites inserted in TRANS in the same mouse (5’ and 3’ loxP sites): ~12% of the projects with at least one positive (targeted) founder mouse.

Two loxP sites inserted in CIS in the same mouse (5’ and 3’ loxP sites): ~25% of the projects with at least one positive (targeted) founder mouse.

 

*KNOCK-IN - LARGE SEQUENCES (Large-KI; STILL UNDER RESEARCH, DEVELOPMENT & OPTIMIZATION): SUCCESS RATE = ~50% (positive knock-in insertion; 5’ and 3’ junction may be altered/missing; random "transgenic" insertions, etc) of the projects with at least one positive (targeted) founder mouse.

Generation of large knock-in (Large KI) mice containing large sequences (e.g. fluorescent proteins, STOP cassettes, IRES-Cre, etc.) usually requires the design of the targeting strategy, production of Cas9 mRNA/protein and 1 or more gRNA, preparation of donor dsDNA/long ssDNA templates, microinjection of one-cell embryos, and screening of founder mice.

 

LEADERSHIP

Jorge HENAO-MEJIA, M.D., Ph.D.; Scientific Director

Leonel D. JOANNAS, LATg; Technical Director

 

STAFF

Matthew MING; BSc. Research Technologist

 

LOCATION

421 Curie Boulevard

Biomedical Research Building II/III, 3rd Floor, Room # 312 (Office) and Room # 320 (Laboratory)

Philadelphia, PA 19104, USA

Contacts

Name Role Phone Email Location
Jorge HENAO-MEJIA, M.D., Ph.D.
Scientific Director CRISPR/Cas9 Mouse Targeting Facility
 
Only e-mail inquiries!
 
jhena@pennmedicine.upenn.edu
 
BRB II/III, 3rd Floor, Room # 312
 
Leonel D. JOANNAS, LATg
Technical Director CRISPR/Cas9 Mouse Targeting Facility
 
Only e-mail inquiries!
 
ljoannas@pennmedicine.upenn.edu
 
BRB II/III, 3rd Floor, Room # 320
 
Matthew MING, BSc.
Research Technologist
 
Only e-mail inquiries!
 
matthew.ming@pennmedicine.upenn.edu
 
BRB II/III, 3rd Floor, Room # 320
 

Map


https://www.google.com/maps/place/Biomedical+Research+Building+II%2FIII,+421+Curie+Blvd,+Philadelphia,+PA+19104/@39.9478702,-75.1990227,17z/data=!3m1!4b1!4m5!3m4!1s0x89c6c65ec9c3306f:0x810d59d739a86378!8m2!3d39.9478702!4d-75.196834