Yale Cancer Center Functional Genomics Core

The YCC Functional Genomics Core provides resources for Yale investigators to perform pooled shRNA and CRISPR/Cas9 screens in human and mouse cells. Resources include pooled libraries for screening, consultation and guidance in performing screens, and bioinformatics support for analysis of screening data. In addition, the core houses genome-wide collections of shRNA constructs and full length human ORF cDNAs that are available as individual constructs.

Contacts:

David A Calderwood

Ben E. Turk

For requests/orders: Clotilde Calderwood

"This Core provided a fantastic service to me. They provided me with a large number of shRNA for different genes within a very short time. We routinely use them in our experiments. I am about to publish one manuscript that describes the use of some of these shRNAs and give the core full credit. The core is just great."

- Ruth Halaban, PhD, Senior Research Scientist in Dermatology

We have obtained genome-wide Mission™ shRNA libraries for human and mouse in the lentiviral pLKO vector. These are stored as arrayed glycerol stocks and contain, on average, 5 hairpins per gene. These shRNA expressing vectors can be provided as individual plasmids (bacterial cultures) or as part of complex pools (plasmid or viral pools).

Individual shRNAs
$44 per clone provided as bacterial culture – discounts available for large orders (please inquire)
$271 per clone provided as lentiviral culture supernatant (1-4 clones)


Premade shRNA pools
$252 per pool provided as pooled plasmid for lentiviral generation.
Please inquire for pricing of pools provided as lentiviral culture supernatant.

Human kinome: 5642 hairpins targeting 523 genes
Human tyrosine kinome: 1198 hairpins targeting 90 genes
Human kinome plus: (protein & lipid kinases plus phosphatases): 5642 protein kinase hairpins (523 genes), 451 lipid kinase hairpins (45 genes), 1556 phosphatase hairpins (249 genes)
Human epigenome: 2013 hairpins targeting 353 genes
Human de-ubiquitinylation: 642 hairpins targeting 102 genes
Human E3 ligases: 3984 hairpins targeting 675 genes
Mouse de-ubiquitinylation: 638 hairpins targeting 101 genes

Pooled shRNA screening:
Pooled screening is not currently offered as a service but consultations are available to allow investigators to perform screens in their own labs, and we can provide protocols for DNA extraction, PCR barcoding, and sample preparation for next generation sequencing.

Bioinformatics analysis of screening data:
Next generation sequencing results of pooled screens performed using our libraries will be mapped onto the input libraries, and enriched or depleted hairpins and genes will be identified using RNAi gene enrichment analysis (RIGER). Mapped normalized read counts and a ranked lists of significantly enriched and depleted genes will be provided as excel files. ($75.48/h - typically will require 3 h/screen but this will vary with complexity – please inquire for more information).

We currently have the following single-vector Addgene CRISPR libraries. These libraries were developed at the Broad Institute and will be distributed under the terms of a UBMTA:

Broad GPP genome-wide Brunello (human single plasmid system, 4 gRNAs for each gene)
Broad GPP kinome Brunello, (human single plasmid system, 4 gRNAs for each gene)
Broad GPP genome-wide Brie, (mouse single plasmid system, 4 gRNAs for each gene)
Broad GPP kinome Brie, (mouse single plasmid system, 4 gRNAs for each gene)

$252 per pool provided as pooled plasmid for lentiviral generation.
Please inquire for pricing of pools provided as lentiviral culture supernatant.


Bioinformatics analysis of screening data:
Next generation sequencing results of pooled screens performed using our libraries will be mapped onto the input libraries, and enriched or depleted hairpins and genes will be identified using RNAi gene enrichment analysis (RIGER). Mapped normalized read counts and a ranked lists of significantly enriched and depleted genes will be provided as excel files. ($75.48/h - typically will require 3 h/screen but this will vary with complexity – please inquire for more information).

We have obtained the ORFeome 1.1, 3.1, 5.1, 7.1, OCAA, OCAB and 8.1 libraries (http://horfdb.dfci.harvard.edu/index.php?page=home), which include full length cDNAs for almost all human genes in a Gateway entry vector. Individual clones are provided as bacterial cultures ($44/clone).

Note that cDNAs from earlier collections (~35% of available clones) are not fully clonal and are not sequence verified. We therefore recommend that investigators streak out cultures and sequence 5-10 colonies to ensure the cDNA is correct.