Massive sequencing of hybrid DNA obtained by DNA:RNA immunoprecipitation (DRIP) in extracellular microvesicles (MV) produced by HeLa cells (using S9.6 antibody)

ABSTRACT: Hybrid (Bound) and non-hybrid (Unbound) DNA from Hela EV was isolated using DRIP procedure as follows. Between 1-5μg of nucleic acid DNA from EVs (DNase0 treated) was incubated overnight with 1μg of anti-DNA-RNA Hybrid (S9.6) antibody (KeraFast) under rotation at 4°C in 500μl of binding buffer (10mM NaPO4 pH 7.0, 140 mM NaCl, 0.05% Triton X-100). The following day, 25μl of A/G magnetic beads (Pierce™) were washed twice in 500μl of binding buffer (30' min each wash) and added for 2h at room temperature under rotation to the S9.6+nucleic acid complex. The nucleic acids bound to the S9.6 antibody (Hybrid fraction or Bound) were separated from those unbound (Non-Hybrid or Unbound) by using a magnetic rack (which captured the Bead+Hybrid fraction). 500μl of the unbound fraction (bead-free) were

INSTRUMENT(S): Illumina MiSeq

ORGANISM(S): Homo sapiens

SUBMITTER: Maurizio Cardelli 

PROVIDER: E-MTAB-9634 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress