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Acta Agronomica Sinica ›› 2023, Vol. 49 ›› Issue (3): 662-671.doi: 10.3724/SP.J.1006.2023.22012

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

Heredity and fine mapping of an early-senescence leaf gene ESL8 in rice

ZHU Xiao-Tong1,2(), YE Ya-Feng2, GUO Jun-Yao2, YANG Hui-Jie2, WANG Zi-Yao1, ZHAN Yue2, WU Yue-Jin2, TAO Liang-Zhi2, MA Bo-Jun1, CHEN Xi-Feng1,*(), LIU Bin-Mei2,*()   

  1. 1College of Chemistry and Life Sciences, Zhejiang Normal University, Jinhua 321004, Zhejiang, China
    2Hefei Institutes of Physical Science, Chinese Academy of Science, Hefei 230031, Anhui, China
  • Received:2022-03-03 Accepted:2022-07-21 Online:2023-03-12 Published:2022-08-19
  • Contact: CHEN Xi-Feng,LIU Bin-Mei E-mail:1634453295@qq.com;xfchen@zjnu.cn;liubm@ipp.ac.cn
  • Supported by:
    Natural Science Foundation of Anhui Province(2108085MC99);Hefei Science and Technology Project(J2020G45);Anhui Science Technology Major Project(202003c08020006);Natural Science Foundation of Zhejiang Province(LD19C130001)

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Abstract:

Early-senescence mutants are genetic materials important for the researches on the molecular mechanism of cell apoptosis in plants. An early-senescence mutant named esl8 (early senescence leaf 8) has been screened from the mutagenesis library of rice variety Kefujing 7. Compared with the wild-type control, the leaves of the esl8 mutant displays a severe of early-senescence phenotype at heading stage in rice, and its agronomic traits, including plant height, tiller number, grain length, grains per panicle, and seed setting rate, were obviously impaired except for the 1000-grain weight. In leaves of the esl8 mutant, the chlorophyll content was abnormally decreased, and the programmed cell death accompanied by excessive accumulation of the reactive oxygen species and the malondialdehyde were detected by histochemical staining. Genetic analysis indicated that the early-senescence phenotype of the esl8 mutant was controlled by a recessive nuclear gene. Based on the strategy of map-based cloning, the esl8 gene was finely mapped into a 359-kb region flanking by two molecular markers (FM12-14 and FM12-15) on chromosome 12. The prediction and verification of candidate genes by PCR sequencing confirmed that esl8 was a new variational allele of the OsSL/ELL1 gene. The sequence of mutation esl8 occurred in the conserved region of the corresponding wild-type gene. Our results provide a theoretical basis for further study on the biofunction and molecular mechanism of the protein encoding by ESL8 in early-senescence process.

Key words: early senescence, rice, esl8, cell apoptosis, mapping and cloning

Table 1

Primers sequences of molecular markers used in fine mapping of rice ESl8 gene"

分子标记
Molecular marker		 正向序列
Forward sequence (5'-3')		 反向序列
Reverse sequence (5'-3')
LSR6 TCTTGCCTCGCTAGGGTTAG CCCACGTTTCTCTTGTCCTC
LSR14 TCCACCACTCTGACGTCTACTAACC CTGCGGGAAGTGTAGGAGAAGC
FM12-1 CCCAGTTTTGCTTAGCCTCTT TTTTGGCATACATCGATTGG
FM12-3 GCCATTTTTAGACGTGGCAAT TTTGCCACTTTTAGTTTTATCGTTT
FM12-7 GGCTGTTGACCATGCTGTAA TGAAGTCCTAGATGCACTTTTTCA
FM12-9 ACATCAGCAGCCATGGTTTT TTTCAACGGTAAAGTCATGTGG
FM12-11 ACATTATGGTTCACGCATGG CGGGACCGTAGGTAAAAAGG
FM12-14 GCCGATGAAACTCCGATAGC TTATCGTGTGAGGGGGAAAG
FM12-15 ACATCCGCGCTTTTCTTTT TAACAAGGGGCAAAGTGGTC

Table 2

Primer sequences for PCR amplification of rice ESL8 gene"

引物名称
Primer name		 正向序列
Forward sequence (5'-3')		 反向序列
Reverse sequence (5'-3')
ESL8-1 TCTTGCCTCGCTAGGGTTAG CCCACGTTTCTCTTGTCCTC
ESL8-2 TCCACCACTCTGACGTCTACTAACC CTGCGGGAAGTGTAGGAGAAGC
ESL8-3 CCCAGTTTTGCTTAGCCTCTT TTTTGGCATACATCGATTGG
ESL8-4 GCCATTTTTAGACGTGGCAAT TTTGCCACTTTTAGTTTTATCGTTT

Fig. 1

Early-senescence phenotype and physicochemical characteristics of rice esl8 mutant A: plant phenotype at seedling stage; B: plant phenotype at heading stage; C: single tiller phenotype at heading stage; D: Trypan blue staining; E: DAB staining; F: NBT staining; G: the measurement of SOD enzyme activity; H: the measurement of MDA content; I: the measurement of chlorophyll content. WT refers to the wide-type control Kefujing 7, and esl8 refers to the early-senescence mutant esl8. Bar: 10 cm; ** represents the significance between the mutant and wild-type control at P < 0.01."

Table 3

Agronomic traits of rice esl8 mutant and the wild-type control"

材料名称
Material name		 株高
Plant height
(cm)		 穂长
Panicle length (cm)		 有效分蘖数
Effective
panicle number		 每穗粒数
Grain number
per panicle		 结实率
Seed-setting
rate (%)		 千粒重
1000-grain
weight (g)
WT 101.18±2.93 14.87±0.86 9.00±1.15 206.00±18.00 81.27±2.18 26.74±0.41
esl8 89.88±1.03** 13.69±0.56** 7.00±0.58** 129.00±10.00** 58.45±2.33** 25.98±0.33

Fig. 2

Fine mapping and sequencing of rice ESL8 gene A: the physical map of the ESL8 gene by preliminary mapping; B: the genetic map of the ESL8 gene by fine mapping; ‘n’ refers to the number of F2 mutants used for mapping; the number under each marker represents the recombinant number detected by the corresponding marker. C: the sequencing results of ESL8 gene in the mutation site. WT refers to the wild-type control Kefujing 7 and ESL8 refers to the early-senescence mutant. The red box indicates the location of ‘TGCCCC’ deletion and ‘GCG’ insertion."

Fig. 3

Sequence alignment and phylogenetic tree of ESL8 proteins A: the sequence alignment of the proteins coding by the esl8 and its wild-type gene; B: phylogenetic tree of ESL8 and its homologous proteins in Arabidopsis and rice; C: the comparison of conserved sequences of ESL8 and its homologous proteins."

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