pCMV-GIN-ZEO Lentiviral shRNAmir

shRNAmir constructs from the human retroviral library have been transferred into the pCMV-GIN-ZEO lentiviral vector developed in the Elledge lab. The pCMV-GIN-ZEO vector has been modified from the pPRIME-CMV-GIN vector (Stegmeier et al 2005) to include a zeocin selectable marker. Unique features of the lentiviral shRNAmir library include:

microRNA-adapted shRNAmir design greatly increases knockdown specificity and efficiency
Efficient single copy knockdown- Important for pooled screens
GFP is a marker of shRNAmir expressing cells
Efficient delivery to primary and non-dividing cell lines
Already cloned into the pCMV-GIN-ZEO lentiviral vector.

Single copy knockdown with pCMV-GIN-ZEO

U2OS cells were transduced with Rb expressing shRNAmir lentiviruses (MOI >0.4). GFP-, dsRed-, and LNGFR-expressing cells were FACS-sorted 4 days after infection, and Neo expressing cells (CMV, CMV-NEO, and CMV-GIN) were selected with 500 ug/ml G418 for 1 week. Drug selection was withdrawn 1.5 days before harvesting the cells. Whole-cell extracts were analyzed by immunoblotting for the indicated proteins. From Stegmeier et al. 2005.

Stegmeier F et al (2005) "A lentiviral microRNA-based system for single-copy polymerase II-regulated RNA interference in mammalian cells" PNAS Vol 102:37; 13212-13217

Silva J et al (2005) "Second-generation shRNA libraries covering the mouse and human genomes" Nature Genetics Vol 37:11, 1281-88.


Vector Element	Utility
CMV Promoter	RNA Polymerase II promoter
cPPT	Central Polypurine tract helps translocation into the nucleus of non-dividing cells
WRE	Enhances the stability and translation of transcripts
GFP	marker to track shRNAmir expression
IRES-NEO	Mammalian selectable marker
AMPr	Ampicillin bacterial selectable marker.
5'LTR	5' long terminal repeat
pUC ori	High copy replication and maintenance of plasmid in e.coli
SIN-LTR	3' Self inactivating long terminal repeat
RRE	Rev response element
ZEO	Bacterial selectable marker

Additional Information:

pCMV-GIN-ZEO Lentiviral shRNAmir

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Send mail to webmaster with questions or comments about this web site.
Copyright © 2005 Gentaur BVBA
Last modified: feb-07