GSM2497587: WT_TAC_W2_48; Mus musculus; RNA-Seq
Cardiomyocytes were isolated using Langendorff perfusion from the left ventricular free wall 2 weeks after the sham operation and 3 days and 1, 2, 4 and 8 weeks after the TAC operation. Echocardiography was used to evaluate left ventricular function and assess whether the heart was appropriately exposed to pressure overload. Rod-shaped live cardiomyocytes (viability of cardiomyocytes was over 80% for all time points) were immediately collected from two mice at each time point with a 0.2–2-µl pipette (sample volume, 0.5 µl) and incubated in lysis buffer. Single-cell cDNA libraries were generated using the Smart-seq2 protocol and the efficiency of reverse transcription was assessed by examining the Ct (Cycle threshold) values from real-time quantitative RT-PCR of control genes (Tnnt2, Cox6a2) using a CFX96 real-time PCR detection system (Bio-Rad) and by examining the distribution of the lengths of cDNA fragments using a LabChip GX (Perkin Elmer) and/or TapeStation 2200 (Agilent Technologies). The following primer sets were used for real-time quantitative RT-PCR: Tnnt2 mRNA forward, TCCTGGCAGA GAGGAGGAAG; Tnnt2 mRNA reverse, TGCAGGTCGA ACTTCTCAGC; Cox6a2 mRNA forward, CGTAGCCCTC TGCTCCCTTA; Cox6a2 mRNA reverse, GGATGCGGAG GTGGTGATAC. A Ct value of 25 was set as threshold. The remaining libraries were sequenced using a Hiseq 2500 system (Illumina).
GEO Accession
GSM2497587