GSM2427897: HEK_pxMARGI_rep1_1; Homo sapiens; OTHER
The major steps of MARGI technologies are: crosslinking, isolation of nuclear material, protein biotinylation, genome fragmentation, RNA fragmentation, stabilization of proteins on streptavidin beads, end modification and RNA ligation, DNA ligation, reverse crosslinking, purification of RNA and DNA, removal of biotin from non-ligated linkers, pulldown of ligation products, reverse transcription, DNA denaturation, circularization and re-linearization, generation of sequencing library, and paired-end sequencing. See Extended Materials and Methods in published paper for details. Sequencing library was generated by amplifying material using NEBnext Hi-fidelity PCR enzyme (New England Biolabs M0541S) with NEBnext multiplex oligos for Illumina sequencing (New England Biolabs E7335S) as the primers. Multiple test PCRs with 1 μL of sample and 10 μL reaction volume were run to identify the correct cycle number for the production run. The production PCR was run in a 50 μL reaction volume with 5 μL sample. After the PCR, size selection was performed using Ampure XP beads (Beckman Coulter A63880). Library between 200-2000 bp was selected. Library was run on a Bioanalyzer to determine the size distribution and then paired-end sequenced for 100 cycles at each end on Illumina HiSeq 2500 (PE100) or MiniSeq sequencers (PE150).