and make the shRNA expression vector for you .
conditions listed on the ordering page. Submitting an order form is construed as
you have carefully read and agree with such terms and conditions.
modified from murine embryonic stem cell virus (pMSCV)
plasmid to express double stranded small hairpin RNA
from human U6 snRNA promoter. A compounded
puromycin resistance-internal ribosome entry site-green
fluorescence protein (puro-IRES-GFP) cassette is inserted
behind the phosphoglycerate kinase (PKG) promoter for
antibiotic selection and green fluorescence labeling in
mammalian cells. Annealed DNA oligos for expression of
small hairpin RNAs are to be cloned behind the human
U6 snRNA promoter between BamHI and Xho I sites. The
small hairpin RNA encoded by pRetro-U6G can be
delivered either by standard plasmid transfection
approach or by infecting target cells with packaged virus
particles. The replication incompetent retroviral particles
generated upon transfection of packaging cells, such as
Phoenix A or E cells, can infect a wide range of target
cells, including hematopoietic and embryonic stem cells,
as well as most other mitotically active cells, and
achieves high level expression of small hairpin RNAs.
All pRetro-U6G derived plasmids should be propagated in
suitable host strains such as DH5a, or HB101, under
selection of resistance to ampicillin (50 mg/ml). E.coli
replication origin: pUC; copy number: high.
Purchase of the shRNA vector Design & Construction service amounts to entering a
service agreement under which Cellogenetics is contractually obligated to:
1. design a requested number of shRNA target sequences based on a set of
widely accepted design rules and proprietary design rules. We generally have 50%
success rate in obtaining shRNA sequences that have 70% percent “knock-down”
efficiency. However, because the functionality of the shRNA target sequences can
only be determined empirically, we offer a "Service Guarantee*" to any purchase
of three or more constructs to be made for each target gene.
2. synthesize the corresponding DNA oligoes for and generate the ordered shRNA
expression constructs.
3. confirm the cloned shRNA coding sequence by DNA sequencing.
4. make glycerol stocks of bacterial clones containing the ordered shRNA
expression constructs and prepare technical spec sheet.
5. ship the ordered shRNA expression constructs as glycerol stocks along with all
pertinent technical information within 15 business days from the date when the
order was placed.
* "Service Guarantee"
Cellogenetics' Service Guarantee applies to any purchase of three shRNA
constructs for each target gene to be designed and constructed by Cellogenetics.
During one year from the date when the constructs were sent, Cellogenetics
guarantees that at least one of the three shRNA constructs will generate 70%
percent “knock-down” or higher efficiency over the target protein or mRNA as either
tested individually or collectively in a pool in transfection experiments. The "knock-
down" efficiency is defined as percentage reduction in expression level of target
protein or mRNA produced from a co-transfected cDNA plasmid in the above
experiment using either protein assay, or functional reporter assay, or RNA assay.
In the event that none of the three constructs surpasses the 70% "knock-down"
efficiency, Cellogenetics agrees to re-design and generates additional shRNA
expression constructs for the same target gene free of charge. For this guarantee,
Cellogenetics reserves the right to be shown reasonable "proof of failure", which
can be the results from any of the above assays or the combination thereof.
