the TRIzol was directly added to the cell sample with the amount 1 mL per 1×10^6 cells, followed by vortex mixing and standing for 5-minute. The mixture was centrifuged at 4°C at 12,000xg for 5 minutes, and the supernatant was transferred to a new EP tube for extraction. 300μl of chloroform/isoamyl alcohol (24:1) was added into the transferred supernatant and the mixture was vigorously vortexed to ensure thorough mixing. Then the mixture was centrifuged at 4°C and 12,000xg for 8 minutes. Take the clear upper aqueous layer and repeat the above process one time. After centrifugation, the final upper aqueous layer  was carefully transferred to a new 1.5ml EP tube with a 2/3 supernatant volume of isopropanol (for low amount of tissues/ cell, 2µL 5mg/mL glycogen can be added for precipitation), gently inverted to mix well, placed in a -20°C refrigerator for 2 hours. Next, the precipitation mixture was centrifuged with 17500xg at 4°C for 25 minutes. The supernatant was discarded, and the precipitation was washed with 0.9ml of 75% ethanol. The precipitation was suspended by inverting the tube up and down several times. Then the precipitation was collected by centrifuging at 17500g for 3 minutes at 4°C and discarding the supernatant. Repeat the centrifugation to remove supernatant completely and the precipitation was dried in the biosafety cabinet for 3-5 minutes. Finally, 20µL~200µL of DEPC-treated or RNase-free water was added to dissolve the RNA. Subsequently, total RNA was qualified and quantified using a Fragment Analyzer or Agilent 2100 Bioanalyzer (Agilent, CA, USA), or Qseq-400 (Bioptic, Taiwan, China). Library preparation is performed using MGIEasy Small RNA Library Prep Kit (BGI-Shenzhen, China). A certain amount of RNA is ligated 3' and 5' adaptors sequentially in two reaction systems for adaptors ligation. After adaptor ligation, RNA is reverse transcribed into cDNA and PCR amplified. The library is fragment size selected by polyacrylamide gel electrophoresis and subjected to quality control. Next, the single-stranded library products are produced via denaturation. The reaction system for circularization is set up to get the single-stranded circularized DNA products. Any single stranded linear DNA molecules will be digested. The final single strand circularized library is amplified with phi29 and rolling circle amplification (RCA) to make DNA nanoball (DNB) which carries more than 300 copies of the initial single stranded circularized library molecule. The DNBs are loaded into the patterned nanoarray and sequencing reads of SE50 bases length are generated on G400 platform (BGI-Shenzhen, China).