title: CRUK/Melton lab-Human melanoma-71-v2-microRNA expression profiling
status: Public on Feb 08 2012
contact.city: Edinburgh
contact.first: Yan
contact.last: Xu
contact.email: kitty2hata@gmail.com
contact.address: Crewe Road South
contact.department: MRC
contact.country: United Kingdom
contact.institute: The University of Edinburgh
organism: Homo sapiens
manufacture_protocol: The MicroRNA Expression Profiling Assay requires an input of only 100-200 ng of isolated total RNA.The input RNA is polyadenylated and converted to cDNA by standard methods. A single miRNA-specific oligo (MSO) is used to assay each miRNA on the panel. All MSOs are hybridized to the sample in parallel, and a solid-phase primer extension step further increases specificity and reduces noise . Universal amplification of extended products only after this point avoids PCRinduced detection bias. The universal PCR step creates fluorescently labeled products identifiable by their unique MSO sequence. These labeled assay products correspond to, and are in relative abundance to, specific original miRNA in the sample. The address sequence from each MSO is used to hybridize specific miRNA products to specific locations on the BeadArrayTM substrate for readout.