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Core Facilities

Collaborative RNAi

Services and Pricing

Researchers are encouraged to contact the Core Coordinator at any stage of the screening process, from assay development to data analysis, for confidential advice on your project and 'navigation' among facilities, resources, etc.

NEW in 2014: Online GESS seed sequence enrichment analysis of RNAi screen datasets. This online tool is relevant to both siRNA and shRNA screens. Post-screen analysis to detect seed sequence enrichment. An important addition to informatics-based approaches to screen validation.

NEW in 2014: Access to genome-scale CRISPR pooled library screening through the Broad Institute Genetic Perturbation Platform (formerly RNAi Platform).

NEW in 2014: Genome Engineering Production Group (GEPG) at HMS provides genome engineering construct services. Useful for development of cell lines for screens as well as for follow-up of screen results. See details below.

Writing a grant that proposes an RNAi or CRISPR screen, or proposes use of genome engineering technologies at large scale? We can also help with advice, key references, budgeting, and letters of support.

Below are listed the various genome engineering services, RNAi reagent libraries, equipment, and related services available from the member groups.

Please note that you can mix-and-match—e.g., get help engineering a custom cell line if that's needed, then use a library from one group and an assay readout instrument at another. In fact, these are exactly the type of interactions we hope to facilitate by joining forces.

*NEW* Support for Genome Engineering

Genome engineering techniques such as CRISPR/Cas-based knockout provides a highly complementary approach that can be used alone, or in combination with RNAi, such as to create a custom cell line for large-scale RNAi screening or confirm RNAi results. Support for Genome Engineering through the DF/HCC Collaborative RNAi Core includes:

(1) Access to genome-wide pooled format CRISPR/Cas9 screening through the Broad Genetic Perturbation Platform (formerly RNAi Platform).

(2) Access to custom CRISPR/Cas9, TALEN and related construct design and build services through the HMS Genome Engineering Production Group, including (a) Custom support for genome engineering is now available through the Genome Engineering Production Group (GEPG) at HMS; and (b) Custom design and construction of CRISPR sgRNAs for knock-out or knock-in, as well as custom donor constructs for knock-in approaches. The GEPG help you harness these exciting new approaches to make mutant cell lines, reporter cell lines, transgenic animals, etc. for small- or large-scale assays. The DF/HCC Collaborative RNAi Core Director (S. Mohr) is available to give presentations on genome engineering to labs and departments.

A. Reagents for Primary Screens (siRNA, shRNA and other Reagent Libraries)

1. Pooled RNAi Screening Libraries—Human or Mouse Cells

2. Arrayed RNAi Screening Libraries—Human Cells

3. Arrayed RNAi Screening Libraries—Mouse Cells

4. Arrayed RNAi Screening Libraries—Drosophila Cells

5. RNAi Screening in whole animals in Drosophila

NEW in 2010: Reagents for miRNA interrogation

B. Reagents for Secondary Screening and Follow-Up

C. Assay Read-Out Equipment

D. Additional Equipment

E. Image Analysis, Statistics & Bioinformatics Support for Screen Data Analysis

F. Cell Lines

G. Additional Reagents & Resources


A. Reagents for Primary Screens (siRNA, shRNA and other Reagent Libraries)

NOTE: Individual shRNA plasmid constructs are available from the DF/HCC DNA Resource Core. Please see their PlasmID website.

Here is an overview of the available reagent sets. See below for details & pricing.

Assay Format

Cells

Coverage

Reagent Type

Center(s)

Pooled

Any Human or Mouse Cell Type

Full-Genome

shRNA (lentivirus)

Various

Arrayed

Transfectable Human and Mouse Cells

Full-Genome and Sub-Sets

siRNA

ICCB-Longwood

Arrayed

Any Human or Mouse Cell Type

Full-Genome and Sub-Sets

shRNA (lentivirus)

Various

Arrayed

Primary & Cultured Drosophila Cells

Full-Genome and Sub-Sets

dsRNA

DRSC

Detailed Information about the Libraries, Assay Read-Out Instruments, and other Services & Equipment Available

New in 2010: Two collections of miRNA mimics and one collection of miRNA inhibitors for human cell screening (through ICCB-L; see their library page).

1. Pooled RNAi Screening Libraries—Human or Mouse Cells

a. Elledge Lab Pooled Libraries. Pooled shRNA libraries described in the following publications are available from Steve Elledge’s lab. Please contact the Core Coordinator or contact the Elledge lab directly for more information.

b. Pooled Mouse or Human Libraries from The RNAi Consortium (TRC). TRC libraries are available in pooled format (in addition to arrayed format; see below) at the DFCI RNAi Facility and the RNAi Platform of the Broad. For more information and pricing, please contact the Core Coordinator or contact a facility directly. Described in: Moffat et al. (2006) A Lentiviral RNAi Library for Human and Mouse Genes Applied to an Arrayed Viral High-Content Screen. Cell. 124(6):1283-1298.

2. Arrayed RNAi Screening Libraries—Human Cells

Arrayed Human siRNA Libraries. Please see ICCB Longwood Screening Facility RNAi Libraries for up-to-date information about the arrayed siRNA libraries available at the ICCB-Longwood Screening Facility. Note that on the ICCB-L website, some siRNA reagents are referred as "pools" because each well in the arrayed library contains a small pool of siRNAs directed against a single target gene.

Arrayed Human shRNA Libraries. Please see The RNAi Consortium (TRC) for an overview of the arrayed TRC shRNA (lentiviral) libraries, which are available at the DFCI RNAi Facility and the RNAi Platform at the Broad Institute.

3. Arrayed RNAi Screening Libraries—Mouse Cells

Arrayed Mouse siRNA Libraries. Please see ICCB Longwood Screening Facility RNAi Libraries for up-to-date information about the arrayed siRNA libraries available at the ICCB-L Screening Facility. Note that on the ICCB-L website, some siRNA reagents are referred as "pools" because each well in the arrayed library contains a small pool of siRNAs directed against a single target gene.

Arrayed Mouse shRNA Libraries. Please see The RNAi Consortium (TRC) for an overview of the arrayed TRC shRNA (lentiviral) libraries, which are available at the DFCI RNAi Facility and the RNAi Platform at the Broad Institute.

4. Arrayed RNAi Screening Libraries—Drosophila Cells

The DRSC has designed, generated and maintains full-genome and additional RNAi libraries for screening (in some cases, in collaboration with other groups). The full-genome library is available for screening on-site at the DRSC and additional libraries are available for screening on- or off-site.

Related information:

DRSC dsRNA Libraries for RNAi Screening
DRSC Library Costs

5. RNAi Screening in whole animals in Drosophila

For information about availability of fly strains for tissue-specific expression of RNAi hairpins (Gal4-UAS system), please see the Transgenic RNAi Project web site.

B. Reagents for Secondary Screening and Follow-Up

After a large-scale primary screen, the next step is usually to verify results using the same and/or additional RNAi reagents directed against the same target genes that "hit" in the primary screen. Doing one to three replicates in the primary screen is typical but doing more replicates is recommended for secondary analysis. In general, the choice of reagents appropriate for secondary analysis reagents will be specific to the library and/or assay used in the primary screen. In some cases, however, it might be appropriate to get reagents from more than one group for follow-up (such as siRNAs from one group and shRNAs from another), particularly if you will switch cell-types as part of your secondary analysis. A number of reagents for follow-up are available from the participating groups. Please contact the core coordinator or the group that provided the library used in the primary screen for more information.

See also, Genome Engineering Production Group, e.g. for follow-up with knockout cell lines.

C. Assay Read-Out Equipment

Instrument

Use

Location*

Molecular Devices Paradigm

Plate-Reader (GFP, FITC, Luciferase)

DRSC (NRB)

PE Packard Fusion

Plate-reader (Fluorescence, Luciferase, FRET, AlphaScreen etc.)

RNAi Platform of the Broad Institute

PerkinElmer EnVision

Plate-Reader (GFP, FITC, Luciferase)

DFCI RNAi Facility

PerkinElmer EnVision

Plate-Reader (GFP, FITC, Luciferase)

ICCB-Longwood (Seeley Mudd)

PerkinElmer EnVision

Plate-Reader (GFP, FITC, Luciferase)

ICCB-Longwood (Seeley Mudd)

PerkinElmer EnVision

Plate-Reader (GFP, FITC, Luciferase, SureFire/Alpha)

ICCB-Longwood (Seeley Mudd)

PerkinElmer EnVision

Plate-Reader (GFP, FITC, Luciferase, SureFire/Alpha)

RNAi Platform of the Broad Institute

MDS SpectraMax

Plate-Reader (190 to 850 nm)

DFCI RNAi Facility

MDS SpectraMax

Plate-Reader

ICCB-Longwood (Seeley Mudd)

Molecular Devices Flexstation III

Plate-Reader with Liquid Handling Automation

ICCB-Longwood (Seeley Mudd)

LiCor Aerius

Far-Red Plate Reader for In-Cell Western

DRSC (NRB)

TTP LabTech Acumen [NEW in 2013]

Laser scanning cytometer (multi-channel)

ICCB-Longwood (Seeley Mudd)

Blueshift Isocyte

Laser scanning cytometer (488 and 532 nm)

ICCB-Longwood (Seeley Mudd)

TTP LabTech Acumen X3

Laser scanning cytometer

RNAi Platform of the Broad Institute

Applied Precision cellWoRx

High-throughput, high-content imaging

ICCB-Longwood

MDS ImageXpress

High-throughput, high-content imaging

ICCB-Longwood (Seeley Mudd)

MDS ImageXpress

High-throughput, high-content imaging

ICCB-Longwood (Seeley Mudd)

PerkinElmer Evotec Opera

High-throughput, high-content confocal imaging

DRSC (NRB)

* Note that purchase, service, etc. for some of the instruments may be costs that are shared among various groups. All require appropriate training and sign-up for use.

D. Additional Equipment

Equipment

Location

Cell Culture Hoods (BL2)

All Locations

Cell Culture Incubators (BL2)

All Locations

Plate Washers & Aspirators

All Locations

Liquid handling Robotics

All Locations

Cell Culture Hoods (BL2+)

DFCI, RNAi Platform at Broad

E. Image Analysis, Statistics & Bioinformatics Support for Screen Data Analysis

We encourage potential screeners to think about data and image analysis early in the process, as simple changes to assay design might have a profound influence on downstream analysis steps.

Image Analysis Support. Most of the screening microscopes (see above) come with software for image analysis. We can also provide access to other software programs and experts. For projects requiring a significant degree of expert input and/or customization, it can be appropriate for these expert(s) to become collaborators (i.e. co-authors). In many cases, screen imaging projects are referred for analysis to Dr. Tiao Xie at the HMS Image Data Analysis Center (IDAC). The IDAC can provide access to open source and commercial software, as well as develop customized solutions when needed.

Statistics & Bioinformatics Support. Many of the centers have database support for screen data that includes basic statistical analysis tools. For example, the ICCB-Longwood ScreenSaver database facilitates data management and analysis for siRNA screens. We can also refer you to specific open source and commercial software packages for statistical analysis and bioinformatics, as well as to expert bioinformaticians who can provide custom support. For projects requiring a significant degree of expert input and/or customization, it can be appropriate for these expert(s) to become collaborators (i.e. co-authors). Please contact the core coordinator for a list of current software packages available at various locations, and/or for an introduction to an appropriate image or data analyst who might be able to help with your project.

See also, GESS online tool for seed sequence enrichment analysis, which can help prioritize hits for more focused follow-up (useful for analysis of large-scale screen data from human or mouse siRNA or shRNA screens, or Drosophila shRNA screens).

The DRSC can provide support for our online COMPLEAT tool for analysis of screen data in the context of protein complexes, including comparison of screen data from more than one condition (e.g. treated versus untreated screen data sets).

The DRSC also developed the UP-TORR tool for up-to-date annotation of the gene-to-RNAi-reagent relationship (fly, worm, mouse and human). UP-TORR is useful to screeners for choosing the right RNAi reagent for a study and for analysis or re-analysis of RNAi data.

As a collective group, the member centers in the DF/HCC Collaborative RNAi Core are interested to provide additional support for screen data analysis and integration. Feel free to contact us about your needs.

F. Cell Lines

Cell lines for Drosophila screening can be requested from the DRSC. We can also help with set-up for large-scale collection of embryos for primary cell screens.

Cell lines for mammalian screening are available from vendors such as ATCC. Please contact the core coordinator for advice on selecting the appropriate cell line(s) for your screen assay.

G. Additional Reagents and Resources

Transfection Reagent Testing Set. Want to transfect siRNAs into your favorite cell line(s) but not sure which transfection reagent is the best for the cell type(s)? The ICCB-Longwood Screening Facility has available a testing set of more than 30 different transfection reagents.

Looking for something else? Please feel free to send us feedback on additional resources, test sets, reagent libraries or other services that the participating groups could provide, either individually or through our collaborative efforts.