|
|
|
|
LibrariesInvitrogen Human 70mer Oligo LibraryThis array was made using the “HEEBO” Human Exonic Evidence Based Oligonucleotide 70mer oligo library from Invitrogen. The set was developed in collaboration with Illumina and researcher at the University of California, San Francisco, Stowers Institute, Stanford University, Rockefeller University, and University of Basel, Switzerland. The 48,958 probe library was generated in part using the software ArrayOligoSelector, an open source tool for selecting oligos. Candidate sequences were filtered based on uniqueness, self-binding, complexity, and GC content. Oligos were preferentially selected based on uniqueness (binding energies >-35kcal/mol for other sequences), 3’ proximity (<1000 based from 3’ end), and the presence of the selected sequence in all transcripts of the gene. The library contains 44,308 probes targeting human genes including constitutive and alternative exonic probes, ESTs, mRNA, BCR/TCR Genic/Regional probes, transgenic/vector probes, viral probes, and microRNA tagged templates. The set also contains a wide array of control sequences, including negative and positive controls. Addition information about the library is available on the Invitrogen website:http://www.invitrogen.com/content.cfm?pageid=11001 This array will be printed on two slides. MGH PGA Mouse 70mer Oligo Library Version 2.0This array was made using the PGA mouse 70mer oligo library generated at Massachusetts General Hospital. The 19,556 oligos were designed using the OligoPicker software generated at the facility and available for free download at http://pga.mgh.harvard.edu/oligopicker/. Oligos were selected using a stringent set of filters to ensure specific representation of all protein-coding sequences in the NCBI GenPept database. The program designs the oligos to have a Tm within a set 10°C range, to ensure hybridization under similar conditions. Also, OligoPicker avoids sequences that are likely to cross-hybridize with other genes or to self-anneal (form primer dimers). The sequence specificity of each oligo is tested with a global BLAST score and all oligos with a BLAST score above a threshold are discarded. Additional information about the library, including sequences of oligos, is available on the MGH website: https://dnacore.mgh.harvard.edu/microarray/index.shtml. Ambion® miRNA Microarray Version 2The Ambion® mirVana™ microRNA probe set includes approximately 500 DNA oligos corresponding to human, rat, and mouse miRNAs. In addition, there are 152 “Ambi-miR” miRNAs; newly identified human miRNAs. This library was generated using sequences available on miRBase Sequence Database version 8.0, the miRNA registry at the Sanger Institute. The amine-modified oligos are 42-46 nucleotides long, including an 18-24 nt segment targeting a specific miRNA. The remaining portion of the oligo includes a linker region to reduce steric hindrance, and an amine modification. The library is spotted onto epoxy coat slides using our OmniGrid 100 array robot. The epoxy surface chemistry of the slide reacts with the amine modification to bind the oligo to the slide. RNA samples used on these arrays must contain a full complement of miRNAs. For this, we recommend isolating RNA with the Ambion® mirVana™ miRNA Isolation kit. This kit ensures that the sample will include smaller species of RNAs. The labeling reaction is performed on RNA that has either been enriched for miRNAs or on the small RNA fraction from gel electrophoresis (this enrichment step is a service that the array facility will provide). miRNA samples are labeled using the Ambion® mirVana™ miRNA labeling kit. This kit utilizes a 3’ tailing reaction that incorporates amine-modified nucleotides. These modified nucleotides are then labeled with fluorescent Cyanine dyes and hybridized overnight on the printed arrays. For more information, please visit the Ambion® website at http://www.ambion.com/techlib/resources/miRNA/index.html. Ambion® mirVana& miRNA probe set printed on Schott-Nexterion® Slide E
| |||||||||
|
