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Acta Agronomica Sinica ›› 2025, Vol. 51 ›› Issue (2): 347-357.doi: 10.3724/SP.J.1006.2025.42030

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Map-based cloning and functional analysis of Dwarf and Tillering 1 (DT1) gene in rice

LI Chun-Mei(), CHEN Jie, LANG Xing-Xuan, ZHUANG Hai-Min, ZHU Jing, DU Zi-Jun, FENG Hao-Tian, JIN Han, ZHU Guo-Lin, LIU Kai()   

  1. Zhongkai University of Agriculture and Engineering, Guangzhou 510225, Guangdong, China
  • Received:2024-06-25 Accepted:2024-09-18 Online:2025-02-12 Published:2024-10-10
  • Contact: E-mail: liukai5088@126.com
  • Supported by:
    State Key Laboratory of Crop Gene Exploration and Utilization in Southwest China(SKL-KF202315);State Key Laboratory for Managing Biotic and Chemical Treats to the Quality and Safety of Agro-products(2021DG700024-KF202408);Basic and Applied Basic Research Foundation of Guangdong Province(2024A1515013028);Science and Technology Plan Project of Yunfu(2023020202);Guangdong Provincial Universities Characteristic Innovation Project(2023KTSCX046);Guangzhou Science and Technology Plan Project(2024A04J4995)

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Abstract:

Tillering is a crucial trait that influences plant architecture and yield in rice. In this study, we identified a natural mutant with dwarf stature and high tillering, which we designated as dwarf and tillering 1 (dt1). The dt1 mutant exhibited significant reductions in panicle length, seed setting rate, grain length, grain width, thousand-grain weight, and the number and size of vascular bundle sheath cells compared to the wild type. Map-based cloning revealed that the dt1 phenotype was caused by an 8 bp insertion in the second exon of D17/HTD1 (LOC_Os04g46470), which encodes Carotenoid Cleavage Dioxygenase 7 (CCD7), a key enzyme in strigolactone biosynthesis. Thus, dt1 represents a new allele of D17/HTD1. Additionally, the dt1 mutant showed significantly reduced germination rate, root length, and root diameter, all of which were restored by the exogenous application of the strigolactone analog GR24. Transcriptomic analysis identified 579 up-regulated and 506 down-regulated genes in the dt1 mutant. Gene Ontology (GO) analysis revealed that the up-regulated genes were significantly enriched in pathways related to auxin response, endogenous stimulus response, and hormone response, while the down-regulated genes were enriched in pathways involved in cellular carbohydrate metabolism and histone methylation. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that the up-regulated genes were associated with plant hormone signal transduction, whereas the down-regulated genes were linked to amino sugar and nucleotide sugar metabolism, as well as diterpenoid biosynthesis. These findings enhance our understanding of the regulatory roles of CCD7 and strigolactones in rice and hold significant theoretical implications for rice breeding.

Key words: rice, dwarf and tillering, map-based cloning, biological function

Table 1

InDel makers used for mapping"

名称Name 正向引物Forward primer (5′-3′) 反向引物Reverse primer (5′-3′)
C2783 ATGGCAAGCACAAGAAGTTGC CGGAGTGGGCATTGTTTTACA
C2782 AACCATTTCACCACAAACCAGC GAGCCTTCGATACTCACCGA
C2763 ACGAGTCCCATCCAAGTACG GAAACAGGAGTAGCTCCCCG
C2754 TTCTTCCGTACAGCAGGCAT AGGAAACTTGAAACGCGCCA

Table 2

Primer sequences used for PCR and cloning"

名称Name 正向引物Forward primer (5′-3′) 反向引物Reverse primer (5′-3′)
OsCCD7-E1 GGAGGCCAAGTCCAAAGATG ACCAGCGTCATATCAGTTCG
OsCCD7-E3 GTGAGTGACGCATGCAGGTT GTCCTTCATGCTCGCAGTGT
OsCCD7-E6 TGTCAAGCTGCTCCTACCAGT CCACAAGTGCATTCTCTGTCC
OsCCD7-E7 GCTCAGTCAGGAGGCTGTC TCCGAGACTGGATGTACAGTAG
OsCCD7-cds CTACAACCTCCTCTTCCGCT AGAAAGTGAAGTGGGATCGC

Fig. 1

Agronomic traits of wild type and dt1 mutant Plant phenotype (A), tiller number (B), plant height (C), panicle phenotype (D), number of grains per panicle (E), number of filled grain per panicle (F), panicle length (G), seed-setting rate (H), 1000-grain weight (I), grain size (J), grain length (K), and grain width (L) of wild type and dt1 mutant at maturity. Bars in A, D and J are 5 cm, 2 cm, and 5 mm, respectively."

Fig. 2

Histological phenotype of the leaves and stems of the wild type (WT) and dt1 mutant at mature stage A: the leaves of the wild type and dt1 mutant at mature stage; B: cross-sections of the wild type leaves at 20× magnification; C: cross-sections of the dt1 mutant leaves at 20× magnification; D: the number of bundle sheath cells of the wild type and dt1 mutant leaves; E: the area of bundle sheath cells of the wild type and dt1 mutant leaves; F: stems of the wild type and dt1 mutant at mature stage; G: cross-sections of the wild type stem at 20× magnification; H: cross-sections of the dt1 mutant stem at 20× magnification; I: the number of bundle sheath cells in the stems of the wild type and dt1 mutant; J: the area of bundle sheath cells in the stems of the wild type and dt1 mutant. Bars in A and F are 2 cm."

Fig. 3

Map-based cloning of rice DT1 gene A: preliminary mapping of DT1; B-C: fine mapping of DT1; D: gene structure of candidate gene LOC_Os04g46470 (black box represents exon, black line represents intron); E: the DNA and amino acid sequences of LOC_Os04g46470 in the WT and dt1 (red represents the mutated DNA or amino acid)."

Fig. 4

Germination of WT and dt1 mutant with or without GR24 treatment A: phenotypes of WT and dt1 mutant germinated for 7 d; B: phenotypes of WT and dt1 mutant germinated for 7 d with GR24 treatment; C: germination rates of WT and dt1 with or without GR24 treatment."

Fig. 5

The root morphology and the results of related parameters of WT and dt1 mutant with or without GR24 treatmentA: root morphology of wild type and dt1 mutant after two weeks of GR24 treatment; B: total root length of wild type and dt1 mutant; C: root diameter of wild type and dt1 mutant; D: root surface area of wild type and dt1 mutant; E: root volume of wild type and dt1 mutant."

Fig. 6

Volcano of differentially expressed genes between WT and dt1 mutant Padj represents the P value adjusted by multiple hypothesis testing. Fold Change represents the fold change of gene expression between WT and dt1. The horizontal and vertical dashed lines indicate the threshold values used for identifying differentially expressed genes, set at -log10 Padj=1.301 and log2 (Fold Change) = 1 or -1, respectively."

Fig. 7

GO and KEGG enrichment analysis of differentially expressed genes A: GO enrichment analysis of differentially expressed genes; B: KEGG enrichment analysis of differentially expressed genes; C: differentially expressed genes in the ABA signaling pathways; D: differentially expressed genes in the GA biosynthesis pathways."

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