Functional Genomics (La Jolla)
The Functional Genomics core facility provides infrastructure for RNAi-based Target Discovery services, from initial feasibility assessment, all the way through verification of identified targets. It also serves as a repository for reagents and assays, expertise and other technical information, and works towards technical improvements when necessary.
The Core’s primary effort is dedicated to loss-of-function screening via RNAi, although gain-of-function screening for specific gene families is also available. In addition it supplies siRNAs, shRNA and ORF clones to aid the screen follow up process.
Our personnel develop both plate reader and image-based cellular assays, and adapt them to HTS-siRNA/shRNA screening conditions in conjunction with the High Content Screening Core. We also collaborate with the Bioinformatics Shared Resource to aid the screening follow-up process.
Consultation: Provide expert advice on all steps in a functional genomics experiment, from initial design to execution of high throughput projects.
Transfection/transduction testing: Evaluate cell line transfectability through a battery of transfection reagents in parallel, both for transfection efficiency and impact on viability. For lentiviral screens, infectivity is assessed.
Assay development: Cellular assays are taken from the researcher, tested under transfection/ transduction conditions, and brought to a 384-well format. Z’ factor is assessed to determine the feasibility of the project. The assay development process is completed by performing a small screen (single 384-well plate) to determine hit frequency.
Screen execution: siRNA/lentiviral collections are plated in 384-well format and screened against plate reader or image based assays. Following QC, raw data are statistically analyzed to identify significant hits. Results are returned to the investigator for hit selection in excel format. siRNAs are performed using individual siRNAs (family focused screens) or RNAi pools (whole genome screening). Screens are performed in duplicate, although there is the option of adding more data points for an additional fee.
Hit cherry-picking and confirmation: Selected hits are cherry-picked and plated in 384-format. Up to 16 hit collection copies are provided to allow a re-run through the primary assay as well as through additional secondary assays. Results are analyzed and returned to the investigator in excel format.
siRNA and lentiviral shRNA or ORF clone distribution: We will provide any siRNA or lentiviral clone present in our libraries at affordable cost.
Custom shRNA set construction and validation: We generate a set of 6 lentiviral-shRNAs (either constitutive or inducible) against any given target and provide 2 ml of viral supernatant of each construct. For an additional fee we will create shRNA cell lines and determine silencing efficiency by QPCR.
The following libraries are available for screening and clone distribution:
- Genome-wide ON TARGET-Plus (OTP) Human siRNA library (Dharmacon): Targets 18,301 genes, both as pools and as four single siRNAs per gene
- Extended human druggable genome siRNA collection (Life Technologies): Targets 7740 genes, four siRNAs per gene
- Human ubiquitinome library (Qiagen): Targets 1170 genes, three siRNAs per gene
- Human kinome library (Life Technologies): Targets 704 genes, four siRNAs per gene
- Human transcriptional regulators library (Life Technologies): Targets 1362 genes, four siRNAs per gene
- Human cancer pathways set (Dharmacon): Targets 252 genes with four pooled OTP siRNAs
- Custom siRNA libraries, either pooled or as single siRNAs.
Lentiviral shRNA libraries:
- Human Kinome Lenti-shRNA library (TRC, SIGMA). Targets 672 genes, averaging 11 shRNA constructs per gene.
- Human miRIDIAN inhibitor library (Dharmacon), targeting 885 miRNAs
- Human miRNA mimic library (Life Technologies),which includes 858 human miRNAs
ORF libraries (gain-of-function screening):
- ORF library, 2440 clones
- Expression-ready lentiviral kinase library, 373 clones
- Expression-ready lentiviral secreted protein library, 858 clones
- Expression-ready lentiviral apoptosis library, 476 clones
- Expression-ready lentiviral innate immunity set, 172 clones
Equipment and Resources
- Benchcell work station with Bravo liquid handling platform (Agilent)
- STAR liquid handling station (Hamilton)
- Tissue culture facility
- WellMate liquid dispenser (Matrix Technologies)
- Micro Flo Liquid Dispenser (BioTek)
- ELx 405 Plate washer (BioTek)
- High-Throughput Microscopes and Plate Readers are utilized via the High Throughput
Screening and High Content Screening
|Assay development plates||plate||$35.00||$43.75||$47.25||$92.05||F319|
|Custom Lentiviral shRNA set- 6 constructs||each||$500.00||$625.00||$675.00||$1,315.00||F105|
|Custom library- OTP pool||each||$8.00||$10.00||$10.80||$21.04||F354|
|Custom library- OTP siRNA/miRNA||each||$6.00||$7.50||$8.10||$15.78||F353|
|Custom library preparation, set up||each||$300.00||$375.00||$405.00||$789.00||F346|
|Custom library- standard siRNA||each||$4.00||$5.00||$5.40||$10.52||F347|
|Hit picking screen follow-up- OTP pool||each||$4.00||$5.00||$5.40||$10.52||F309|
|Hit picking screen follow-up- OTP siRNA/miRNA||each||$3.00||$3.75||$4.05||$7.89||F308|
|Hit picking screen follow-up -standard siRNA||each||$3.00||$3.75||$4.05||$7.89||F311|
|Hs. Cancer pathways pooled OTP_screen||duplicate set||$200.00||$250.00||$270.00||$526.00||F102|
|Hs. Druggable V3 (4x)_screen||duplicate set||$4,000.00||$5,000.00||$5,400.00||$10,520.00||F332|
|Hs. Kinase (4x)_screen||duplicate set||$750.00||$937.50||$1,012.50||$1,972.50||F221|
|Hs. Lenti-ORF_Apoptosis_screen||duplicate set||$800.00||$1,000.00||$1,080.00||enquire||F101|
|Hs. Lenti-ORF_extracellular proteins_screen||duplicate set||$1,200.00||$1,500.00||$1,620.00||enquire||F301|
|Hs. Lenti-ORF_kinome_screen||duplicate set||$800.00||$1,000.00||$1,080.00||enquire||F300|
|Hs. Lenti-shRNA Kinome (4x)_screen||duplicate set||$5,000.00||$6,250.00||$6,750.00||enquire||F103|
|Hs. miRNA inhibitors_screen||duplicate set||$600.00||$750.00||$810.00||$1,578.00||F343|
|Hs. miRNA mimics_screen||duplicate set||$600.00||$750.00||$810.00||$1,578.00||F342|
|Hs. OTP pool miniscreen_random set_screen||duplicate set||$300.00||$375.00||$405.00||$789.00||F104|
|Hs. T. Factors (4x)_screen||duplicate set||$1,000.00||$1,250.00||$1,350.00||$2,630.00||F294|
|Hs. Ubiquinome (3x)_screen||duplicate set||$1,000.00||$1,250.00||$1,350.00||$2,630.00||F328|
|Hs. Whole Genome OTP (4x)_screen||duplicate set||$25,000.00||$31,250.00||$33,750.00||$65,750.00||F349|
|Hs. Whole Genome OTP pooled library_screen||duplicate set||$6,000.00||$7,500.00||$8,100.00||$15,780.00||F350|
|lentiviral ORF clone (LB)||each||$50.00||$62.50||$67.50||enquire||F128|
|Lentiviral sup-2ml prep||each||$50.00||$62.50||$67.50||$131.50||F129|
|Non Specific siRNA control (1nmol, 20uM)||each||$35.00||$43.75||$47.25||$92.05||F312|
|ORF clone-entry vector (LB)||each||$20.00||$25.00||$27.00||$52.60||F127|
|Other Products (other consumables)||each||$1.00||$1.25||$1.35||$2.63||F355|
|Other Services (consulting, labor etc.)||min||$0.84||$1.05||$1.13||$2.21||F356|
|Plasticware, disposables, various||each||$50.00||$62.50||$67.50||$131.50||F326|
|single set- Hs. miRNA mimics 1X||single set||$300.00||$375.00||$405.00||$789.00||F335|
|single set-Hs. Druggable V3 4x||single set||$2,000.00||$2,500.00||$2,700.00||$5,260.00||F154|
|single set-Hs. Kinase 4x||single set||$375.00||$468.75||$506.25||$986.25||F124|
|single set-Hs. miRNA inhibitors 1X||single set||$300.00||$375.00||$405.00||$789.00||F336|
|single set-Hs. T. Factors 4x||single set||$500.00||$625.00||$675.00||$1,315.00||F194|
|single set-Hs. Ubiquinome 3x||single set||$500.00||$625.00||$675.00||$1,315.00||F117|
|single set-Hs. Whole Genome OTP 4x||single set||$12,500.00||$15,625.00||$16,875.00||$32,875.00||F351|
|single set-Hs. Whole Genome OTP pooled library||single set||$3,000.00||$3,750.00||$4,050.00||$7,890.00||F352|