关键词: 黄瓜, 远红光, 弱光胁迫, 混池测序, FRS转录因子, 单核苷酸多态性

Abstract:

【Objective】Cucumber is the most widely cultivated greenhouse vegetable in China. However, low light stress frequently occurs during protected cultivation, which often induces excessive stem elongation and spindly growth of cucumber plants. Screening for cucumber germplasm with reduced susceptibility to spindly growth under low light stress and identifying the underlying genes can provide a theoretical basis for genetic improvement of low light tolerance in cucumber. 【Method】Using a low light stress screening system, cucumber germplasm was evaluated. Subsequently, a segregating population was constructed. The Bulked Segregant Analysis sequencing (BSA-seq) method was employed to identify significant loci associated with the target trait. Functional annotation and gene family analysis were then performed to elucidate candidate genes involved in low light stress response. 【Result】Two distinct cucumber materials, designated as long hypocotyl (CS ) and short hypocotyl (CR), were initially identified. Light quality treatment assays indicated that the differential response of hypocotyl between CS and CR was primarily associated with far-red light perception. Using CS and CR as parental lines, F₁ and F₂ populations were developed. Genetic analysis of hypocotyl length under low light stress revealed that the trait was predominantly governed by two novel QTL loci—LSH1 (Chr.4) and LSH2 (Chr.5). Through bulked segregant analysis (BSA), these loci were predicted to encompass 373 and 163 candidate genes, respectively. Subsequently, key components of the far-red light signaling pathway were systematically identified in cucumber, including Phytochrome A (PHYA), FAR-RED ELONGATED HYPOCOTYL 1 (FHY1)/FHY1-LIKE (FHL), and FAR1 RELATED SEQUENCE (FRS) transcription factors. The analysis yielded two PHYA genes (CsPHYA1 and CsPHYA2) on chromosome 6, one CsFHY1 gene on chromosome 3, and 22 CsFRS transcription factor genes distributed across all chromosomes except chromosome 2. Notably, CsFRS12 was physically located within the LSH1 interval on chromosome 4. Phylogenetic and motif analyses indicated that CsFRS12 contained three conserved domains: a FAR1 DNA-binding domain, a MULE transposase domain, and a SWIM Zinc finger domain. CsFRS12 clustered within subfamily Ⅰ and showed closest homology to Arabidopsis AtFRS2 (AT2G32250), AtFAR1 (AT4G15090), and AtFHY3 (AT3G22170), all key regulators of far-red light signaling. Cloning and sequence comparison revealed multiple SNPs between CsFRS12^CR and CsFRS12^CS alleles, which were significantly correlated with hypocotyl length under low light stress. Yeast transcriptional activity assays demonstrated that CsFRS12^CR possessed significantly lower transcriptional activity than CsFRS12^CS. Expression profiling further indicated that CsFRS12 was highly expressed in the stem and hypocotyl tissues of cucumber. In summary, these findings highlight CsFRS12 as a promising candidate gene underlying low light-induced hypocotyl elongation in cucumber. 【Conclusion】Two cucumber materials with differential responses to low light stress were identified, and two significant loci (LSH1 and LSH2) associated with hypocotyl elongation under low light stress were mapped using BSA-seq. Furthermore, within the LSH1 interval, a key transcription factor CsFRS12, which participates in the far-red light signaling pathway, was found as a critical candidate gene for low light stress response in cucumber.

Key words: cucumber, far-red light, low light stress, BSA, FRS transcription factor, SNP