2.11 | RT-PCR
According to the manufacturer’s instructions, total RNA was extracted from LF tissues and cells using the Trizol reagent (Invitrogen) and converted to cDNA using the Reverse Transcription Synthesis Kit (Takara, Dalian, China). Then, the gene expression levels were analyzed using a Thermal Cycler Dice Real-Time system (Takara, China). After the processes of reaction, the threshold cycle (Ct) was determined and realtive mRNA level was normalized to the GAPDH expression using the 2-∆Ct method. Designed primers and their sequences were all summarized in Table 2 and Table 3.