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Fig. 1 | Molecular Horticulture

Fig. 1

From: A domestication-associated gene, CsLH, encodes a phytochrome B protein that regulates hypocotyl elongation in cucumber

Fig. 1

(A) Hypocotyls of 10-d-old wild type (LH) and long hypocotyl (lh) mutant seedlings under dark conditions. Bar, 5 cm. (B) Hypocotyls of 10-d-old LH and lh cucumberseedlings under white light conditions. Bar, 5 cm. (C) Hypocotyl length of the indicated genotypes in (A) and (B). Error bars indicate ± SD (n = 20). The asterisk indicates a significant difference (P < 0.05, t test). (D) Maximum likelihood tree of phytochrome protein sequences from Arabidopsis thaliana (At), tomato (Solanum lycopersicum; Sl), and cucumber (Cucumis sativus; Cs). The sequence data were deduced from amino acid sequences taken from the Arabidopsis Information Resource (https://www.arabidopsis.org/), Tomato Genome Database (https://solgenomics.net/), and Cucurbit Genome Database (http://cucurbitgenomics.org/). (E) The 7-bp deletion detected in the first exon of CsaV3_3G015190 in the lh mutant; this deletion results in amino acid changes and a premature stop codon. (F) Genotyping of 2000 F2 individuals using two CsLH flanking markers: SNP_chr03_11192318 and SNP_chr03_11332009. L = homozygous SNP alleles from Chinese Long 9930 (reference genome); l = homozygous SNP alleles from lh; H = heterozygous SNP alleles. (G) CsLH rescued the long hypocotyl phenotype exhibited by the lh mutant at 10 days old. Bar, 5 cm. plh: CsLH = CsLH coding sequence driven by the lh promoter. (H) Hypocotyl lengths of the genotypes indicated in (G). Error bars indicate ± SD (n = 20). Different lowercase letters (a, b) indicate significantly different hypocotyl lengths in pairwise tests (P < 0.05, Tukey’s test). (I) Semi-quantitative RT- PCR analyses of CsLH expression in lh, LH, and plh: CsLH. The internal reference was the α-TUBULIN (TUA) gene. The primers for testing CsLH expression were designed based on the 7-bp deletion sequence; therefore, no band that could be amplified from the lh sequence. (J) Nucleotide diversity of the CsLH region in five cucumber groups, as measured using the π value. Error bars indicate ± SD. EA = East Asian, EU = Eurasian, IN = Indian cultivated and wild genotypes; IND = Indian Domestic, INW = Indian wild genotypes, XSBN = Xishuangbanna. (K) Matrix depicting unique and shared SNPs among the EA, EU, XSBN, and IN (including INW and IND) groups at the CsLH region. The yellow bars indicate total SNP number of each cucumber group. (L) Nucleotide diversity of five cucumber populations at individual nucleotide sites in CsLH. The x axis indicates the genome position in the Chinese Long 9930 v2 sequence, while the y axis indicates the π value of each cucumber group. (M) Pairwise difference of allele frequency (FST). The x axis indicates the genome position in Chinese Long 9930 v2, while the y axis indicates FST value of INW vs. IND, INW vs. EA, INW vs. EU, and INW vs. XSBN. (N) Number of haplotypes observed in each cucumber group. Green background highlight that XSBN only contained H2. (O) and (P) Representative EA (O) and EU (P) seedlings displayed longer hypocotyls than XSBN grown under white light conditions. Seedlings were 14 days old. Bar, 5 cm. (Q) Hypocotyl lengths of the indicated genotypes in (O) and (P). Error bars indicate ± SD (n = 20). Different lowercase letters indicate groups of significance difference in pairwise tests (P < 0.05). (O′, P′) Comparison of hypocotyl lengths among 7-d-old EA, EU, and XSBN seedlings grown under dark conditions. Bar, 1 cm. (Q’) Hypocotyl lengths of the indicated genotypes in (O′) and (P′). Error bars indicate ± SD (n = 20). Lowercase letters indicate no significant differences in the hypocotyl lengths of dark-grown EA, EU, and XSBN seedlings (P < 0.05). (R) Relative expression of CsLH under light and dark conditions in 9930, XIS48, and LH. Different lowercase letters indicate a significant difference (P < 0.05, Tukey’s test, three replications). (S) Model of the genome of the introgression line XSBN 3.2, as genotyped using SNP markers. Green and blue indicate background of parental line 9930 and XIS48, respectively. (T) Hypocotyl lengths of 5-d-old Chinese Long 9930 and XSBN3.2 seedlings grown under white light. Bar, 1 cm. (U) Hypocotyl lengths of indicated genotypes in (T). Error bars indicate ± SD (n = 20). Asterisk indicates a significant difference (P < 0.05)

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