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Integrated analysis in bi-parental and natural populations reveals CsCLAVATA3 (CsCLV3) underlying carpel number variations in cucumber

Abstract

Key message

Carpel number variation in cucumber was controlled by a single gene, Cn . Linkage and association analysis revealed CsCLV3 as the candidate gene of the Cn locus.

Abstract

Carpel number (CN) is an important fruit quality trait of cucumber, but the genetic basis of CN variations is largely unknown. In the present study, segregating analysis in multiple bi-parental mapping populations (F2, F3, and RILs) derived from WI2757 (CN = 3) × True Lemon (CN = 5) suggested that CN is controlled by a simply inherited gene, Cn, with CN = 3 being incompletely dominant to CN = 5. Initial linkage mapping located Cn in a 1.9-Mb region of cucumber chromosome 1. Exploration of DNA sequence variations in this region with in silico bulked segregant analysis among eight re-sequenced lines allowed delimiting the Cn locus to a 16-kb region with five predicted genes including CsCLV3, a homolog of the Arabidopsis gene CLAVATA3. Fine genetic mapping in F2 and RIL populations and association analysis in natural populations confirmed CsCLV3 as the candidate gene for Cn, which was further evidenced from gene expression analysis and microscopic examination of floral meristem size in the two parent lines. This study highlights the importance of integrated use of linkage and association analysis as well as next-gen high-throughput sequencing in mapping and cloning genes that are difficult in accurate genotyping. The results provide new insights into the genetic control of CN variations in cucumber, which were discussed in the context of the well-characterized CLAVATA pathway for stem cell homeostasis and regulation of meristem sizes in plants. The associations of carpel number with fruit shape, size, and weight in cucumber and melon are also discussed.

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Acknowledgments

The authors thank Kristin Haider for technical help and language editing of the manuscript. SL’s work in YW’s lab was partially funded by Shanxi Scholarship Council of China. TB’s was supported by the Fulbright Program. This research was supported by the USDA-CRIS project 3655-21000-048-00D and a U.S. Department of Agriculture (USDA)-Specialty Crop Research Initiative grant (SCRI, project# 2011-51181-30661) to YW. Names are necessary to report factually on available data; however, the USDA neither guarantees nor warrants the standard of the product, and the use of the name by USDA implies no approval of the product to the exclusion of others that may also be suitable. USDA is an equal opportunity provider and employer.

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Correspondence to Yiqun Weng.

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The authors declare no conflict of interest.

Additional information

S. Li and Y. Pan contributed equally to this work.

Communicated by R. Toepfer.

Electronic supplementary material

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Supplementary material 1 Table S1 Information of all markers used in genetic mapping and candidate gene expression studies of carpel number (Cn) locus. Table S2 Distribution of SNPs in 123 re-sequenced cucumber line in 16 kb region harboring the Cn locus. Only SNPs in coding regions are shown. Only the fourth SNP in the first gene (CsCLV3 at 271311 position of scaffold03078, highlighted in red) shows the consistent alleles associated with carpel numbers in all examined lines.   Fig. S1 A. Carpel numbers of two parental lines, True Lemon (CN=5) and WI2757 (CN=3), and their F1 (CN=3). B. Representative fruits of RIL plants from WI2757 × True Lemon showing variations of carpel numbers. Fig. S2 QTL analysis of carpel number in the recombinant inbred line population of WI2757 × True Lemon. A. Frequency distribution of carpel number in 91 RILs based on CN mean of each RIL in 2014 field trial. B and C. QTL mapping with SLAF marker data identified only one QTL for carpel number in this RIL population in cucumber chromosome 1. D. The peak location of LOD curve is consistent with the Cn position when the data was used in linkage analysis with CN as a qualitative trait. Additional online materials File 1 WI2757 and True Lemon CsCLV3 candidate gene sequence alignment with CLUSTAL2.1 (green: SNPs between WI2757 and True Lemon; red: critical SNP differentiating multiple CN=3 and CN=5 lines in association analysis; yellow: 156 bp exon/CDS). (PDF 2395 kb)

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Li, S., Pan, Y., Wen, C. et al. Integrated analysis in bi-parental and natural populations reveals CsCLAVATA3 (CsCLV3) underlying carpel number variations in cucumber. Theor Appl Genet 129, 1007–1022 (2016). https://doi.org/10.1007/s00122-016-2679-1

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Keywords

  • Melon
  • Shoot Apical Meristem
  • Floral Meristem
  • Bulk Segregant Analysis
  • Fruit Shape