title: GSM7051503: 61_1048_C57BL/6NCrl_F; Mus musculus; RNA-Seq
library_name: GSM7051503
library_construction_protocol: Whole kidney total RNA were extracted using Qiagen's RNeasy Mini Kit. Sample 1- 67 and sample 98 - 188 were submitted to the Genome Access Technology Center at the McDonnell Genome Institute at University of Washington in St. Louis. Samples 1- 67, sample 98 - 145 were prepared according to Clontech SMARTer library kit manufacturer's protocol, sample 146 - 188 were prepared according to polyA selection library kit manufacturer's protocol. Sample 1- 67 were indexed, pooled, and sequenced on Illumina HiSeq2500 platform for 50-bp single-end. Sample 98 - 188 were indexed, pooled, and sequenced on llumina NovaSeq S4 2X150 (4 lanes). Sample 68 to Sample 97 were indexed, pooled and sequenced at BGI genomics with DNBseq platform according to the manufacturer's protocol. For sample 146 - 188, Total RNA integrity was determined using Agilent Bioanalyzer or 4200 Tapestation.  Library preparation was performed with 5 to 10ug of total RNA with a Bioanalyzer RIN score greater than 8.0. Ribosomal RNA was removed by poly-A selection using Oligo-dT beads (mRNA Direct kit, Life Technologies). mRNA was then fragmented in reverse transcriptase buffer and heating to 94 degrees for 8 minutes. mRNA was reverse transcribed to yield cDNA using SuperScript III RT enzyme (Life Technologies, per manufacturer's instructions) and random hexamers. A second strand reaction was performed to yield ds-cDNA. cDNA was blunt ended, had an A base added to the 3' ends, and then had Illumina sequencing adapters ligated to the ends. Ligated fragments were then amplified for 12-15 cycles using primers incorporating unique dual index tags. Fragments were sequenced on an Illumina NovaSeq-6000 using paired end reads extending 150 bases.Basecalls and demultiplexing were performed with Illumina's bcl2fastq software with a maximum of one mismatch in the indexing read.