Serveis i Equipaments Principals Functional Genomics
The Functional Genomics Core (FGC) of the Institute for Research in Biomedicine (IRB Barcelona, Catalunya, Spain) is open to work with researchers from academia and industry. We provide comprehensive support to your projects whenever genome-wide measurements of DNA or RNA are needed or gene expression should be manipulated.
Tools for Genomic Analysis
FGC provides tools for:
- miRNAs
- transcriptomics
- DNA copy numbers
- ChIP-Seq
- genomic sequencing
- SNPs
Genome-wide measurements of DNA alterations and gene expression on the RNA level can be a challenge during research projects. FGC provides a complete service to work with you throughout the entire project. In addition to cutting-edge tools like microarrays from Affymetrix and Agilent and Next Generation Sequencing by Illumina , FGC is ready to help with all questions related to your genomic analysis project.
Alteration of Gene Expression
FGC provides:
- human shRNAs
- mouse shRNAs
- human ORF clones
Reverse genetics needs efficient tools to alter gene expression. FGC provides close to 200,000 clones for genetic manipulation. These include all of Sigma's TRC1 human and mouse shRNA clones for knock-down of expression, together with Open Biosystems human ORF clones for over-expression. ORF clones also come handy as probes for in situ hybridization or northerns.
shRNA clones come in lentivirus vectors for efficient infection of many cell types, including non-dividing ones.
ORF clones contain the Gateway system for hassle-free transfer into any kind of expression vector.
Tools for Genomic Analysis & Alteration of Gene Expression
Tools for Genomic Analysis
Which type of genomic analysis tools works best for answering a specific question is dependent on the nature of the question. In general, questions regarding know and well-characterized transcripts (mRNAs and miRNAs) are answered more straight forward using microarrays. Gene discovery and characterization of ChIP material is done more efficiently by sequencing. Some frequently interrogated themes are described here:
RNA
microRNA analysis
FGC provides the Affymetrix miRNA GeneChip for miRNA analysis. For over 70 organisms, a comprehensive analysis of known miRNAs is performed on a single array. Minimal amount: 250 ng total RNA
Discovery of novel small RNAs (miRNAs, piRNAs and other types of small RNAs) is most efficiently performed by sequencing. Minimal amount: 500 ng total RNA
mRNA expression profiling
For over 20 organisms, we provide Affymetrix expression arrays for mRNA profiling. These arrays provide a robust and reliable tool to quantify expression of the majority of known genes. Minimal amount: 5 ng
For over 100 organisms, we provide Agilent expression arrays for mRNA profiling. These arrays are especially suited for small genomes since multiple samples can be analyzed on a single array, leading to cost reduction. Minimal amount: 25 ng total RNA
RNA Seq
By nature, this is performed by sequencing. It allows characterization of alternative splicing, transcriptional start and stop sites and discovery of novel transcripts. Sample processing and data analysis are highly experimental and do not follow standard operating procedures yet. Minimal amount: 100 ng total RNA
DNA
DNA copy number variation (CNV analysis)
CNV analysis at a resolution of 10-100kb is performed on all organisms where expression arrays are available from Affymetrix or Agilent. Minimal amount: 500 ng genomic DNA Especially for organisms with a smaller genome (drosophila, yeast etc.), CNV analysis is also performed by genome sequencing. Together with CNV information, many other types of genetic alterations (point mutations, InDels, translocations, inversions) are studied in the same experiment. Minimal amount: 100 ng genomic DNA
ChIP-Seq
Characterization of transcription factor binding sites and location of histone modifications is best performed by sequencing the ChIP material. In organisms with a complex genome like human or mouse, the optimal baseline (control antibodies, ChIP-input) is not determined yet. Minimal amount: 10 ng ChIPed DNA
DNA methylation
Analysis of multiple loci for DNA methylation is still performed in an experimental phase. For specific projects regarding DNA methylation, please contact FGC.
Alteration of Gene Expression
Reverse genetics needs efficient tools to alter gene expression. FGC provides close to 200,000 clones for genetic manipulation. These include all Sigma's TRC1 shRNA clones for human and mouse for knock-down of expression and Open Biosystems' human ORF clones for over-expression. ORF clones also come handy as probes for in situ hybridization or northerns.
Human & mouse shRNAs
shRNA clones come in lentivirus vectors for efficient infection of many cell types, including non-dividing ones. On average, we provide 4-5 different clones per gene. In many cases, the different clones targeting the same gene provide different knock-down efficiencies and help to control off-target effects. Approximately 75% of all human and mouse refseq genes are targeted. The complete list of shRNA clones is available here. Detailed information about the TRC1 clones is available on Sigma's RNAi webpage.
Human ORF clones
The Open Biosystems ORF library provides approximately 13,000 clones for expression of one open reading frame per gene. These clones are most frequently used for over-expression of genes but can also be used as probes. ORF clones contain the Gateway sytem for hassle-free transfer into any kind of expression vector.
The list of available ORF clones is available here. Detailed information about the ORF clones is available on Open Biosystem's ORF webpage.
General
Initial consultation
Prior to starting your experiments, we invite you to discuss the experimental design with us.
Requirements for the services
Every service (clone delivery, microarray analysis, sequencing...) has its own specifications; details are available on the order forms of the specific services.
Who can perform projects with the FGC?
FGC is a facility open to everybody, researchers at IRB, in Barcelona, Spain, Europe and the rest of the world. We work with academia as well as with industry.
How is payment of services organised at the FGC?
In general, we have two prices, one for IRB researchers and one for everybody else.
Please find here the list of prices as well as the orders forms for all services:
2. Order forms for IRB Barcelona users:
3. Order forms for external users:
For Spanish researchers: The government still does not give research institutions a tax-exempt status. Therefore, you have to add IVA to our prices.
Tools for Genomic Analysis
How long does it take until I get my results?
For smaller microarray projects (
For sequencing projects, a time frame of 3-6 weeks can be given for raw data delivery.
Which organisms can be analyzed?
In the field of Next Generation Sequencing, a reference genome of the organism must be available. Sequences are mapped to this reference genome.
Any organisms with a well annotated genome (or transcriptome) can be analyzed. In the field of microarrays, standard arrays can be purchased for many organisms. Affymetrix covers over 20 organisms, Agilent over 100. Arrays can also be customized for your favorite organism. This is usually easier with Agilent than with Affymetrix.
How do I best prepare my samples?
In general, a combination of organic phase extraction (phenol) with column purification (e.g. Qiagen) works best. These two completely different purification principles remove all kind of impurities. Protocols can be found and downloaded here. For miRNA analysis, do not use columns. Almost all of them remove short RNAs.
For very small samples (a few thousand cells), we propose a Picoprofiling service combining a specific protocol for RNA extraction with robust cDNA amplification method (See Pico-profiling order form).
I have doubts about the quality of my samples. Should I still submit them for analysis?
Before we start any expensive experiment like arrays or sequencing, we perform a broad range of quality controls. These include Nanodrop testing of purity, RNA integrity test on the Bioanalyzer, DNA or RNA specific quantification using the Qubit assays. If we have doubts about the samples, we will contact you for your decision "No or Go".
Alteration of Gene Expression
Who can request clones?
IRB has a single-use contract with Sigma and Open Biosystems, i.e. only IRB researchers and IBMB-CSIC researchers have access to the Sigma libraries; only IRB researchers have access to the Open Biosystems library. If you work outside of IRB, please contact Sigma and IRB administration to find out if these contracts can be modified to include your institution.
For which genes do you have shRNA or ORF clones?
Our shRNA clones cover approximately 75% of human and mouse refseq mRNAs, the ORF clones cover the same percentage of human genes. Access to Clones libraries files to find and download the complete lists of available clones and corresponding genes.
I cannot open the clone lists because they are in xlsx-format.
The libraries contain more than 65,535 clones. Therefore, one needs Excel 2007 to show more than 65,535 rows.
I cannot find my favorite gene in your clone lists.
Our lists contain the official gene names as available on the NCBI website. At NCBI, go to the search field, click on gene and check if you used the official name.
How long does it take until I receive my clones?
Send us an email with your clone request, we tell you when tubes for bacterial inoculation should be delivered and you will receive the clones the same day of tube delivery. In toto, this takes 3 days max.
Propers esdeveniments
