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Acta Agronomica Sinica ›› 2022, Vol. 48 ›› Issue (6): 1389-1400.doi: 10.3724/SP.J.1006.2022.12035


Phenotypic characterization and gene mapping of an early senescence leaf H5(esl-H5) mutant in rice (Oryza sativa L.)

ZHENG Chong-Ke1(), ZHOU Guan-Hua1, NIU Shu-Lin1,2, HE Ya-Nan1, SUN wei1, XIE Xian-Zhi1,*()   

  1. 1Shandong Rice Research Institute / Shandong Academy of Agricultural Sciences, Jinan 250100 Shandong, China
    2College of Life Sciences, Shandong Normal University, Jinan 250014, Shandong, China
  • Received:2021-05-27 Accepted:2021-10-19 Online:2022-06-12 Published:2021-12-05
  • Contact: XIE Xian-Zhi E-mail:zhengck1983@163.com;xzhxie2010@163.com
  • Supported by:
    Fund:The study was supported by the Strategic Priority Research Program of Chinese Academy of Sciences(XDA24030101-6)


A stable mutant esl-H5 (early senescence leaf H5) was identified from the mutant library of japonica rice Huaidao 5 population induced by ethyl methane sulfonate (EMS) treatment. The mutant was normal at seedling stage. However, the lower leaves in esl-H5 mutant displayed premature senescence at about 50 days after sowing. Compared with the wild type (WT), the heading date of the esl-H5 mutant was delayed, while agronomical traits including plant height, panicle length, grain number per panicle, effective tiller numbers, and 1000-grain weight were significantly reduced. Moreover, chlorophyll content was also decreased in esl-H5 mutant. Genetic analysis indicated that the early senescence trait in esl-H5 mutant was controlled by a single recessive gene. ESL-H5 gene was localized on chromosome 1 using molecular marker. MutMap analysis further revealed that one nucleotide G to A replace occurred in the last exon of Os01g0533000 gene which encodes callose synthase. The G to A replace in the ESL-H5 introduced a premature stop codon. Phylogenetic analysis showed that ESL-H5 was homology with Arabidopsis AtGSL7 (Glucan Synthase-Like 7). At tillering stage, the contents of soluble sugar and starch were significantly increased in the leaves of the esl-H5 mutant compared with those of the WT. These results implied that the mutation of ESL-H5 affected the transport of photosynthetic products, resulting in premature leaf senescence phenotypes. The qRT-PCR analysis revealed that the expression levels of disease resistance-related genes PR1a, PR1b, PR2, PR4, PR5, and PR10 in esl-H5 mutant were higher than those in WT, which was consistent with the observation that esl-H5 mutant improved bacterial blight resistance. The present results lay the foundation for studying the roles of sugar signal in regulating rice senescence and disease resistance.

Key words: rice (Oryza sativa L.), early senescence, gene mapping, callose synthase

Table 1

Primers for qRT-PCR"

Primer name
Forward sequence (5'-3')
Reverse sequence (5'-3')

Fig. 1

Phenotypes of esl-H5 mutant A: phenotypes after being sowed for 10 d, bar: 1 cm; B: plant phenotypes at tillering stage, bar: 5 cm; C: plant phenotypes at filling stage, bar: 5 cm; D: phenotypes of upper three leaves at heading stage, bar: 5 cm; E: spike phenotypes at mature stage, bar: 1 cm; F and G: phenotypes of grain length and grain width, bar: 1 cm."

Table 2

Comparison of agronomic trains between esl-H5 mutant and wild type"

Agronomic trait
esl-H5 野生型
Wild type
抽穗期 Heading date 124.20 ± 0.28** 120.80 ± 0.23
株高 Plant height (cm) 60.80 ± 0.42** 87.95 ± 0.62
分蘖数 Tiller number 5.75 ± 0.32** 11.65 ± 0.42
穗长Main panicle length (cm) 15.04 ± 0.23** 15.23 ± 0.21
每穗实粒数 Grain number per panicle 64.50 ± 2.65** 158.60 ± 4.80
一级枝梗数No. of primary branch 12.28 ± 0.25** 13.00 ± 0.37
二级枝梗数No. of secondary branch 20.08 ± 0.86** 27.00 ± 1.23
粒长 Grain length (mm) 7.82 ± 0.25 7.84 ± 0.21
粒宽Grain width (mm) 3.76 ± 0.17** 3.88 ± 0.11
千粒重 1000-grain weight (g) 20.90 ± 0.01** 27.20 ± 0.01

Fig. 2

Relative expression levels of senescence and photosynthesis related genes in WT and esl-H5 mutant A: relative expression levels of senescence related genes in WT and esl-H5 mutant; B: relative expression levels of photosynthesis related genes in WT and esl-H5 mutant."

Fig. 3

Chlorophyll content in esl -H5 mutant and wild type at seedling and tillering stages A: the chlorophyll content of the newly developed leaves after being sowed for 40 days; B: the chlorophyll content of the second leaves from the uppermost after being sowed for 40 days; C: the chlorophyll content of the newly developed leaves after being sowed for 60 days; D: the chlorophyll content of the second leaves from the uppermost after being sowed for 60 days; **: P < 0.01; *: P < 0.05."

Table 3

Separation ratio of three F2 segregation population"

Cross combinations
F1表型 F1 phenotype F2表型 F2 phenotype
Wild type
Wild type
χ2 (3:1)测验
χ2 (3:1) text
WT/esl-H5 15 0 109 35 0.01
NPB/esl-H5 23 0 672 217 0.14
9311/esl-H5 19 0 105 31 0.24

Table 4

Information of markers displayed polymorphism between the pools"

Marker name
Primer sequence (5'-3')
Enzymes used

Fig. 4

Mapping and cloning of ESL-H5 genes A: fine mapping of ESL-H5 (the marker above the horizontal line represents the InDel or SNP marker used in fine mapping, and the number below represents the number of recombinants); B: changes of G to A in the genome of Os01g0533000; C: SNP3 is converted to dCAPS marker; D: plants with early senescence phenotype was detected using dCAPS markers. E: relative expression level of Os01g0533000 in WT and esl-H5."

Fig. 5

Protein structure prediction and phylogenetic tree analysis of ESL-H5 A: protein structure prediction of wild-type and esl-H5 mutant, VTA1, FKS1, and Glucan synthase refer to protein domains; B: ESL-H5 transmembrane region prediction; C: phylogenetic tree analysis of callose synthase in Arabidopsis and rice; D: alignment of amino acid sequences of ESL-H5 and AtGSL7."

Fig. 6

Relative expression patterns of ESL-H5 genes"

Fig. 7

Starch and soluble sugar contents in wild type and mutant leaves at tillering stage A: soluble sugar contents at 6:00; B: soluble sugar contents at 18:00; C: starch contents at 6:00; D: starch contents at 18:00. *: P < 0.05; **: P < 0.01."

Fig. 8

Identification of resistance to bacterial blight of wild type and mutant at tillering stage A: phenotype of bacterial blight resistance; B: statistics of lesion length; C: relative expression levels of pathogenesis-related protein. **: P < 0.01."

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