*4.2. Growth Conditions of Plants in the Field*

All materials were grown in plots at the Baguazhou base of Nanjing Agricultural University in Nanjing, Jiangsu. Nanjing is located in a subtropical monsoon climate zone. The pH of the soil is 6.5, and chemical properties included 0.91 g/kg total N content; 18.91 mg/kg available phosphorus (P) content; 185.67 mg/kg exchangeable potassium (K) and 11.56 g/kg organic matter. We applied Ca(H2PO4)2, 30 kg P/ha and KCl, 60 kg K/ha as basal applications to the plots three days before transplanting. We use urea as N fertilizer, and applied 40% before transplanting, 30% at tilling, and 40% before the heading stage. Plots size was 2 × 2.5 m and the seedlings planted in a 10 × 10 array [32,59–62]. We randomly chose five seedlings from each plot, avoiding those on the edges. The agronomic characters of plant height, total tiller number per plant, panicle length, seed setting rate per plant, grain weight per panicle, grain number per panicle and yield per plant were measured at the maturity stage. Plant height indicated the height of the highest panicle. One panicle from each plant was chosen for calculating the panicle length, grain weight per panicle and grain number per panicle [33]. The agronomic traits of the samples are listed in Supplementary Tables S2 and S3.

#### *4.3. Gene Expression Analysis*

Total RNA was extracted from leaf tissue using TRIzol reagent (Vazyme Biotech Co, Ltd., People's Republic of China). DNase I-treated total RNAs were subjected to reverse transcription (RT) with HiScript II Q Select RT SuperMix for qPCR (+gDNA wiper) kit (Vazyme Biotech Co, Ltd., People's Republic of China) according to the manufacturer's instructions. Quantitative assays were performed using AceQTM qPCR SYBR Green Master Mix (Vazyme Biotech Co, Ltd., China). Relative expression level is normalized to the amount of *OsActin* (LOC\_Os03g50885) in the same sample and presented as 2–CT. All primers used for RT-qPCR are listed in Supplementary Table S6.

### *4.4. T-DNA Insertion Loci Analysis*

Leaf tissues harvested from the plants at 10 days were grown in water. Genomic DNA isolation was performed using the CTAB extraction procedure [63]. T-DNA insertion loci of two overexpression transgenic lines were determined by TAIL-PCR following the procedures previously described [64]. The primers are listed in Supplementary Table S5.
