GSM4514015
GSM4514015: crac_Nanos2CTL_Rep1_NNNTGGAGC; Mus musculus; OTHER
After UV-crosslinking, lysates were prepared from SSCs, and protein-RNA complexes were isolated with antibody, protease cleavage and Nickel-affinity purification, under denaturing conditions. RNA-protein complexes were eluted from Nickel-columns and separated by SDS-PAGE and transferred to a membrane. Proteins were digested and RNA was extracted. Cross-linked RNA molecules were partially digested with RNase A and T1 and subjected to phosphatase treatment. A 3' barcode, which is complementary to the reverse transcription primer, is ligated to RNA. A 5' barcode, which includes the experimental barcode and the UMI, was ligated to RNA. After protein digestion, RNA was extracted and reverse transcribed. cDNA was PCR amplified for 28 cycles using primers compatible with Illumina sequencing. DNA fragments of 130-190 bases were extracted and purified from an agarose gel.
GEO
GSM4514015
GEO Accession
GSM4514015