GSM6704941: F: adjacent normal lung tissue; Homo sapiens; RNA-Seq
The tissue Dissociation Kit (Tumor Dissociation Kit, Human) is used. Mechanical Dissociation and enzymatic hydrolysis are carried out according to the Dissociation operation steps. The cell suspension is first filtered through 70um and then filtered through 30um, if the number of red blood cells in the single cell suspension is large, Red blood cells need to be removed with RBC lysate to prepare a single cell suspension.After the tissue samples were made into single-cell suspension, luna-FL automatic cell counting apparatus was used to count and measure cell viability. The activity of cultured cells was higher than 90% (the activity of primary cells was higher than 70%). If the cell activity could not meet the standard, the dead cells should be removed with the desiccated cell kit to improve cell viability. The cell concentration was adjusted to the ideal concentration of 700-1200 cells /μL After the amplification products were qualified, the sequencing library was constructed.The cDNA is first chemically broken to 200 to 300bp   The left and right fragments, cDNA fragmentation, terminal repair and addition of A, cDNA fragment screening, P7 Adaptor connector connection and PCR amplification to introduce sample Index, finally the fragment screening to obtain the cDNA library.