title: GSM8498388: Mouse TTFs WT 2; Mus musculus; RNA-Seq
library_construction_protocol: Mice were euthanized and tail tendon fascicles were immediately dissected, placed in 2mL Eppendorf Lock tubes, snap frozen in liquid nitrogen and stored at -80C. On the day of isolation, tubes were placed immediately in dry ice. Samples were transferred to supercooled Spex microvial cylinder (6757C3, SPEX™ SamplePrep) containing 150µl of GENEzol™ reagent (GZR200, Geneaid). Samples were cryogrinded in a bath of liquid nitrogen using FreezerMill (6870, SPEX™ SamplePrep) until fascicles were completely disrupted. Pulverized samples were collected in a 1.5 ml tube by rinsing the tubes with 900µl GENEzol™ and kept on ice. To purify RNA, tissue homogenate was mixed well with 200µl of Chloroform (102445, Merck) at a mixing ratio of 1:5. Samples were then spun down for 15min (at 15,000g, 4°C). The upper RNA-containing aqueous phase was transferred to a clean 1.5 ml tube and mixed with one part of 70% ethanol. RNA cleanup was subsequently performed using RNeasy Plus Micro Kit (74034, Qiagen), including a step to remove genomic DNA (Qiagen gDNA Eliminator spin columns). Cleaned RNA concentration and quality were determined by spectrophotometric determination of A260/A280 ratio. Samples were stored at -80 °C, and only samples with A260/A280 ratio of 1.8-2 were used. RNA libraries were prepared for sequencing using standard Illumina protocols