2.3 RNA isolation, cDNA cloning, and sequence analysis
Total RNA from leaves of C. sinensis (SCZ) was isolated using a
Fast Pure Plant total RNA Isolation Kit (Vazyme Cat, RC401-01, Nanjing,
China) according to the manufacturer’s instructions. The cDNA was
synthesized by reverse transcription of the total RNA using Prime Script
RT Master Mix (Vazyme, China). The open reading frame sequences were
amplified using Phusion ® High-Fidelity DNA Polymerase (New England
Biolabs, Ipswich, MA, USA); primers for the clonedCsTPS1/1 -AS gene are shown in Table S1. The PCR products
were purified using a Gel Extraction Kit (CWBIO, Jiangsu, China). The
resulting target cDNA fragment was ligated into the pGEX-4T1 vector and
then subsequently transformed into Trans 1-T1 competent cells.