title: GSM6722290: Tumor_1; Homo sapiens; RNA-Seq
library_construction_protocol: 1 Around 100mg fresh sample or flash frozen tissue in liquid nitrogen is taken for next step.  2 The tissue homogenizer is inserted on ice. Add the 2 mL pre-chilled Homogeniza Buffer.  3 Transfer minced tissue to the chilled tissue homogenizer for 3-5 mins until the tissue is fully infiltrated.  4 Grind the tissue with the tissue homogenizer for around 10-15 times until grinding forces disappear. Assess if the nuclei are completely released into suspension and observe the suspension background under microscopy observation. 70 µm and 40 µm cell strainers are used to filter the tissue homogenate, followed by collection of into a 15 ml centrifuge tube.  5 Centrifuge at 500g and 4℃ for 5 mins and discard the supernatant. The cell pellets are washed twice with Wash buffer.  6 Resuspend the precipitate with the desired volume of Blocking Buffer and mix thoroughly. Observe the nuclei suspension background under microscopy observation.   7 If the suspension background is clean, cell counting can be used. Otherwise, fluorescence-activated cell sorting is needed to filter the suspension background. 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.