Transcriptome analysis of mib1-3 mutant

Shoot buds (2 weeks after germination) and the young pods (2 days after pollination) of WT and mib1-3 mutants were collected with three biological replicates. RNA was extracted by the RNA Kit R6827-01 (Omega, Shanghai, China). We performed RNA-seq using the Illumina HiSeq X Ten platform (Illumina, San Diego, California, USA). The raw sequences were submitted to the NCBI SRA database with accession numbers SRR16944233–SRR16944244. Number of reads per kilobase of exon region in a gene per million mapped reads (RPKM) was used to value expression levels (Mortazavi et al., 2008 (link)), and VC1973A version 1.0 was used as the reference genome (Kang et al., 2014 (link)). Based on the methods described by Audic and Claverie (1997) (link), DEGs were identified. Heat maps were generated by the pheatmap package (https://cran.r-project.org).
For qRT-PCR, the first strand cDNA was synthesized via Takara PrimeScript™ RT reagent Kit RR047A (TaKaRa, Dalian, China). qRT-PCR analysis was conducted using TB Green™ Premix Ex™ RR420A (TaKaRa) and the ABI StepOnePlus machine (Applied Biosystems, Foster City, CA, USA). Three biological replicates with three technical repeats were conducted.