GSM1311876: supernatant_adult_testis_rep1_RNAseq; Mus musculus; RNA-Seq
Chromatoid body was purified according (Meikar et al. 2010 Exp Cell Res). Briefly after dissection seminiferouse tubels were dissociated with collagenese to the cell suspension.  The cells were crosslinked with formaledehyed, resuspend in RIPA buffer and sonicated.  The resulting lysates were centrifuged and supernatant or pellet1 were collected. The pellet1 was resuspended in RIPA buffer, sonicated again and filtrated through 5ul filters.  After centrifugation pellet2 was resuspended in RIPA buffer and incubated with Dynabeads-IgG (lnvitrogen) bounded either with rabbit MVH, IgG or ACR antibodies. After washes the IP complexes were reversed cross-links in RIPA and RNA was extracted with Trizol reagent (Invitrogen). total RNA was prepared with Trizol reagent (Invitrogen) Transcriptome libraries were prepared according  (Armour et al. 2009, Nat Methods) using not-so-random priming (NSR). 1 ug total RNA was revere transcribed using SuperScript III enzyme with first-strand NSR primer. RNA template was removed with RNase H (Invitrogen) Then cDNA was mixed with exo− Klenow fragment (NEB) second-strand NSR primer to synthesize the second strand. For PCR amplification, purified second-strand synthesis reaction were mixed with ExpandPLUS enzyme (Roche) and PE-P5-SBS3 and PE-P7-SBS8 primers. Products were run on a 2% low-melt agarose gel, and the 350–500 bp. range were purified.
GEO Accession
GSM1311876