title: BGISEQ-500 sequencing; Trancriptomic profiling of HCT116 colon cancer cells with or without PBRM1 knock-down by RNAi
design: Trancriptomic profiling of HCT116 colon cancer cells with or without PBRM1 knock-down by RNAi
library_construction_protocol: HCT116 cells were infected with lentivirus containing non-targeting shRNA or shRNA targeting PBRM1. After 72h of infection, cells were harvested and total RNA was extracted. Alternatively, HCT116 cells were transfected with non-targeting siRNA or siRNA targeting PBRM1. After 72h of transfection, cells were harvested and total RNA was extracted. Total RNA was extracted from HCT116 cells using RNeasy kit from Qiagen according to manufacturer's instructions  Oligo (dT) magnetic beads are used to select mRNA with polyA tail. Fragment the target RNA and reverse transcription to double-strand cDNA (dscDNA) by N6 random primer. End repair the dscDNA with phosphate at 5' end and stickiness 'A' at 3' end, then ligate adaptor with stickiness 'T' at 3' end to the dscDNA. Two specific primers are used to amplify the ligation product. Denature the PCR product by heat and the single strand DNA is cyclized by splint oligo and DNA ligase.
attributes.tag: Experimental Factor: Non-targeting siRNA
attributes.value: rna interference