Fig. 4
Association test and expression profile of NOG1 variation. a Sequences comparison of a promoter region between SIL176 and Guichao 2. b Association testing with grain number per main panicle of 16 variants in the 4.3-kb fine-mapping region. Black dots represent 16 variations. c Triangle matrix of pairwise linkage disequilibrium. d Effect of the 12-bp insertion and 15 SNPs in NOG1 promoter by transient expression assays. Above, constructs of pGreenII 0800-LUC vector. Left, constructs with site-directed NOG1 promoter. SIL176 and Guichao 2 define the constructs with the ~2500-bp promoter fragments from SIL176 and Guichao 2, respectively. The SIL176 promoter contained one copy of the 12-bp and the 15 SNPs-type 1 (ACTTCTAATCCAGAG). The Guichao 2 promoter contained two copies of the 12-bp and the 15 SNPs-type 2 (TGCCTAGCCTTGTTA). A1, A2, and A3 define constructs with site-directed mutations at the 12-bp InDel (positions −2214) and 15 SNPs (positions from −2166 to −405) in the SIL176 promoter fragment, respectively. A4 and A5 define constructs with site-directed mutations at TBOXATGAPB and DOFCOREZM TFBS on the 12-bp InDel in the SIL176 promoter fragment, respectively. Right, expression levels of firefly luciferase relative to Renilla luciferase (n = 10). e Comparison of NOG1 gene expression levels between varieties containing one copy of 12-bp and varieties containing two copies of 12-bp (n = 20). Data are means, with error bars showing SEM (two-tailed Student’s t test; **P < 0.01)
