Synthesis kit

Total RNA was extracted using TRIzol reagent (Invitrogen, USA), and then synthesized into first-strand cDNA using a synthesis kit (Thermo Fisher, USA). Gene expression levels were subsequently detected by the Applied Biosystems 7500 real-time PCR system (Thermo Fisher, USA) using the SYBR Green master mix (Thermo Fisher, USA). Relative expression levels were calculated using the 2^−ΔΔCt method. Each sample was performed in triplicate.

Total RNA was isolated by TRIzol reagent (Invitrogen, USA) and the concentration of total RNA was detected by NanoDrop spectrophotometer (Thermo Fisher Scientific, USA). CDNA was synthesized by Synthesis Kit (Thermo Fisher Scientific, USA), and SYBGreen I Master Mix (Roche, Germany) was used for qRT-PCR. 18 S was used as an endogenous control. All data were normalized to internal control, and relative expression was calculated by the 2–ΔΔCT method. Hsa_circ_0005397 primers were purchased from Guangzhou (Geneseed, China), and other primers were purchased from Shanghai (Sangon Biotech, China). The primers used in this study were as follows: hsa_circ_0005397 (Forward: 5′-GACAAAGACAGCA GGTTCCT-3′; Reverse: 5′-CTC​TGT​TCT​GCT​TCT​GAG​TA-3′); EIF4A3 (Forward: 5’-CAGGGCGTGTTTTTGATATGAT-3’; Reverse: 5’-ATCAGCTTCATCCAAAACCAAC-3’; RHOT1 (Forward: 5’-CTGATTTCTGCAGGAAACACAA-3’; Reverse: 5’-GCAAAAACAGTAGCACCAAAAC-3’); and 18 S rRNA (Forward: 5’-GTAACCCGTT GAACCCCATT-3’; Reverse: 5’-CCATCCAATCGGTAGTAGCG-3). After qRT-PCR, PCR products were examined by 2% agarose gel electrophoresis. The experiments were performed at least three times independently with triplicate samples.

Total RNA was isolated with TRIzol reagent (Thermo Fisher Scientific). First-strand complementary DNA (cDNA) was obtained with a Synthesis Kit (Thermo Fisher Scientific); 2 μL of total cDNA and Synergy Brands (SYBR) Green PCR Master Mix (Applied Biosystems) were mixed. Then Eppendorf Master Cycle Realplex2 was used for real-time PCR (40 cycles). The RT-qPCR conditions were as follows: 3 min of enzyme activation at 95 °C, followed by denaturation at 95 °C for 20 s and annealing of primers at 60 °C for 20 s, and extension at 72 °C for 20 s. qPCR data were analyzed by the ΔΔCT method.

An RNeasy Plus Micro Kit (Qiagen, Germany) was used to extract the RNA. Ten oocytes were prepared for each sample. cDNA was produced by using a synthesis kit (Thermo, US), and Gapdh was used as the reference gene. The primer sequences were as follows: Akr1b3 forward: 5′-AACGTGATACCTAGTGACACCG-3′; reverse: 5′-AGCCAGGTTTGTTCAAGATCC-3′; Gapdh forward: 5′-GGTTGTCTCCTGCGACTTCA-3′; reverse: TGGTCCAGGGTTTCTTACTCC. A real-time PCR kit was used (Takara, US). The relative gene expression was calculated by the ΔΔCT method.

The total RNA was extracted using a TRIZOL reagent (Sigma, UK). Total RNA was reverse-transcribed into cDNA with synthesis kit (Thermo, K1671, US) according to manufacturer's instructions. Power SYBR Green PCR Master Mix (Life Technologies, 4368706, USA) in StepOnePlus Real-Time PCR System (Applied Biosystems, USA) was used for qPCR analysis. RPL41 expression was used as internal control. TXNIP, E-cadherin, and Vimentin expressions were normalized to RPL41 using the 2−ΔCt method. Primers used in qPCR reactions are presented in Supplementary Table 1.

Total RNA was extracted from rice seedlings or specific tissues at reproductive stage using Trizol according to the manufacturer's instructions (Invitrogen, Shanghai, USA). Approximately 1 μg of total RNA was used for first-strand cDNA synthesis with the Synthesis Kit (Thermo Fisher Scientific, USA). The reaction products were diluted ten-fold and used as the template for real-time PCR with three biological replicates using the SYBR Premix Ex Taq kit (Takara, Dalian, China). qRT-PCR was performed with an Applied Biosystems Quant Studio 5 (Thermo Fisher Scientific, USA) real-time PCR System. The gene-specific primers used in the qRT-PCR analysis are listed in Table S3.