GSM4721827: Wilms' tumor primary culture CF-00108; Homo sapiens; RNA-Seq
Total RNA was extracted at BGI and analyzed on a Agilent 2100. Library was constructed using the BGISeq-500 Transcriptome library construction protocol: 1. Calculate the DNA input and prepare the library adapters. 2. End-repair step following Carøe et al. 2017 with the addition of 1 μl "Reaction enhancer" consisting of 25% Poly-Ethylene Glycol (PEG4000); 2 μg/μL Bovine Serum Albumin (BSA) and 400 mM NaCl. 3. Ligation step following Carøe et al. 2017 by using 1 μl 10 μM BGISeq-500 adapters. 4. Fill-in step of the adapter to complete library building and purification using 1:5 volume of PB binding buffer (Qiagen) and using Monarch DNA Cleanup Columns (New England Biolabs, Massachusetts, USA). 5. Wash with 750 µl buffer PE (Qiagen). 6. Incubate for 5 minutes at 37 °C. 7. Elute in 40 µl buffer EB (Qiagen). 8. To determine the number of cycles used in index PCR, performe a qPCR in a 20 µl reaction volume using 1:20 dilution of: purified library template, 0.2 mM dNTPs (Invitrogen), 0.04 U/µl AmpliTaq Gold DNA polymerase (Applied Biosystems, Foster City, California, USA), 2.5 mM MgCl2 (Applied Biosystems), 1X GeneAmp, 10X PCR Buffer II (Applied Biosystems), 1 µl SYBR Green (Invitrogen, Carlsbad, California, USA), 0.2 µM forward and reverse primers mixture(CommonprimerBGI forward primer and one of the indexed reverse primers), 13.48 µl AccuGene molecular biology water (Lonza) 
GEO Accession
GSM4721827