For the gene expression assay, total RNA extraction and cDNA synthesis were performed using an RNA extraction kit (TaKaRa, Kusatsu, Japan) and a PrimeScript RT reagent kit (TaKaRa), respectively. Then, quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) was carried out using the SYBR Premix Ex Taq kit (TaKaRa) with an Applied Biosystems 7500 real-time PCR system (Applied Biosystems, CA, United States). The tomato EF-1α gene served as the reference gene71 (link). Each qRT-PCR assay was repeated with three biological samples. The relative expression levels of genes were calculated using the 2–ΔΔCt method.
For the mature miRNA159 expression analysis, the cDNA was generated through reverse transcription from the total RNA by using the miRcute miRNA First-Strand cDNA Synthesis Kit (TIANGEN, Beijing, China). The qRT-PCR analysis was performed using an miRNA qPCR Detection Kit (SYBR Green) (TIANGEN). The tomato U6 small nuclear RNA gene was used as an internal control. The gene-specific primers used in these procedures are listed in Supplementary TableS4 .
For the mature miRNA159 expression analysis, the cDNA was generated through reverse transcription from the total RNA by using the miRcute miRNA First-Strand cDNA Synthesis Kit (TIANGEN, Beijing, China). The qRT-PCR analysis was performed using an miRNA qPCR Detection Kit (SYBR Green) (TIANGEN). The tomato U6 small nuclear RNA gene was used as an internal control. The gene-specific primers used in these procedures are listed in Supplementary Table
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