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Precise flowering timing is critical for the plant life cycle. Here, we examined the molecular mechanisms and regulatory network associated with flowering in Chinese cabbage ( Brassica rapa  L.) by comparative transcriptome profiling of two Chinese cabbage inbred lines, “4004” (early bolting) and “50” (late bolting). RNA-Seq and quantitative reverse transcription PCR (qPCR) analyses showed that two positive nitric oxide (NO) signaling regulator genes,  nitrite reductase  ( BrNIR ) and  nitrate reductase  ( BrNIA ), were up-regulated in line “50” with or without vernalization. In agreement with the transcription analysis, the shoots in line “50” had substantially higher nitrogen levels than those in “4004”. Upon vernalization, the flowering repressor gene  Circadian 1  ( BrCIR1 ) was significantly up-regulated in line “50”, whereas the flowering enhancer genes named  SUPPRESSOR OF OVEREXPRESSION OF CONSTANCE 1  homologs ( BrSOC1s ) were substantially up-regulated in line “4004”. CRISPR/Cas9-mediated mutagenesis in Chinese cabbage demonstrated that the  BrSOC1-1/1-2/1-3  genes were involved in late flowering, and their expression was mutually exclusive with that of the nitrogen signaling genes. Thus, we identified two flowering mechanisms in Chinese cabbage: a reciprocal negative feedback loop between nitrogen signaling genes ( BrNIA1  and  BrNIR1 ) and  BrSOC1s  to control flowering time and positive feedback control of the expression of  BrSOC1s .

international journal of molecular sciences

Nitrogen Signaling Genes and SOC1 Determine the Flowering Time in a Reciprocal Negative Feedback Loop in Chinese Cabbage (Brassica rapa L.) Based on CRISPR/Cas9-Mediated Mutagenesis of Multiple BrSOC1 Homologs