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Acta Agronomica Sinica ›› 2025, Vol. 51 ›› Issue (4): 1110-1117.doi: 10.3724/SP.J.1006.2025.42042

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Phenotypic analysis and gene mapping of a floury endosperm mutant we2 in rice

ZHU Jian-Ping1,2,3(), LI Wen-Qi1,2,3, XU Yang1,2,3, WANG Fang-Quan1,2,3, LI Xia1,2,3, JIANG Yan-Jie1,2,3, FAN Fang-Jun1,2,3, TAO Ya-Jun1,2,3, CHEN Zhi-Hui1,2,3, WU Ying-Ying1,2,3, YANG Jie1,2,3,*()   

  1. 1Institute of Food Crops, Jiangsu Academy of Agricultural Sciences / Key Laboratory of Germplasm Innovation in Downstream of Huaihe River (Nanjing), Ministry of Agricultural and Rural Affairs / Nanjing Branch of Chinese National Center for Rice Improvement, Nanjing 210014, Jiangsu, China
    2Zhongshan Biological Breeding Laboratory, Nanjing 210014, Jiangsu, China
    3Jiangsu Co-Innovation Center for Modern Production Technology of Grain Crops, Yangzhou University, Yangzhou 225009, Jiangsu, China
  • Received:2024-09-07 Accepted:2024-12-12 Online:2025-04-12 Published:2024-12-27
  • Contact: E-mail: yangjie168@aliyun.com
  • Supported by:
    National Natural Science Foundation of China(32201861);Jiangsu Provincial Key Research and Development Program(BE2023362);Project of Seeds Innovation in Jiangsu(JBGS (2021) 008)

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Abstract:

A floury endosperm mutant, we2 (white endosperm2), was identified from a 60Co-irradiated mutant pool of the rice (Oryza sativa) variety Nipponbare. The phenotype and physicochemical properties of we2 were analyzed, and an F2 population derived from a cross between we2 and the indica variety 9311 was used for fine mapping of the target gene. Compared with the wild type, we2 exhibited a white endosperm phenotype with irregularly shaped, loosely packed compound starch granules. The 1000-grain weight, total starch content, and amylose content in we2 were significantly lower than those in the wild type, whereas the lipid content was higher. Genetic analysis revealed that the floury endosperm phenotype of we2 is controlled by a single nuclear recessive gene. For map-based cloning, the we2 mutant was crossed with 9311, and F2 individuals were analyzed. The WE2 locus was initially mapped to chromosome 6 and subsequently fine-mapped to a 244 kb genomic region containing 27 predicted open reading frames (ORFs). Quantitative real-time PCR (qRT-PCR) analysis demonstrated that the expression levels of several genes involved in starch biosynthesis were reduced in the we2 mutant. This study provides a foundation for the cloning and functional characterization of WE2, contributing to a deeper understanding of the genetic and molecular mechanisms underlying rice endosperm development.

Key words: rice, endosperm, starch, fine mapping

Table 1

Primers for fine mapping of WE2"

名称
Name		 正向引物
Forward primer (5′-3′)		 反向引物
Reverse primer (5′-3′)
P22 GACGACACGCACAGCATC GGGGGTTTATATTGATTTTTGCGA
P27 GGACATACCTAGCGCTAGC AGTACTCTGATCACCGTCAGT
P31 CGAGGACGACAGAACGAAGA CGTGTTGAGCCCCACCAG
P36 TGTTTGGAGAGAGTTGGAGATT GGATCGGTTTTCACAAACAGAC
P38 AACAATTCACTCGCCACAGC CGGCGGTTTTGGATTTGGAT
P39 AGCACAGTACTATCGACGGG GGCCATTTTGTGCTGGTACA

Fig. 1

Phenotypic analysis A and B: seeds of WT and we2. C and D: cross-sections of seeds of WT and we2. E: grain length, n = 20. F: grain width, n = 20. G: grain thickness, n = 20. Bars: 1 mm. Values are means ± SD, using Student’s t-test. ** indicates P < 0.01."

Fig. 2

Agronomic traits analysis A-D: 1000-grain weight, plant height, No. of tillers per plant and seed setting rate, n = 20. Values are means ± SD, using Student’s t-test. ** indicates P < 0.01."

Fig. 3

Physicochemical properties analysis A-D: starch content, amylose content, lipid content and protein content, n = 3. Values are means ± SD, using Student’s t-test. ** indicates P < 0.01."

Fig. 4

Scanning electron microscopic analysis Observation of cross-sections of seeds of WT (A-C) and we2 (D-F). A, D: bars: 1 mm; B, E: bars: 50 μm; C, F: bars: 10 μm."

Table 2

Genetic analysis"

杂交组合
Cross combination		 野生型种子数
No. of wild type seeds		 突变体种子数
No. of mutant seeds		 χ2(3:1)
we2/wild type F2 127 40 0.050
Wild type/we2 F2 148 55 0.369

Fig. 5

Fine mapping of WE2 The WE2 was mapped to a 244 kb region with markers P38 and P39 on the long arm of chromosome 6, which contains 27 candidate genes."

Table 3

Candidate genes for WE2"

开放阅读框
ORFs		 基因登录号
Locus ID		 功能注释
Functional description
ORF1 Os06g0659400 Protein of unknown function DUF231, plant domain containing protein
ORF2 Os06g0659500 Similar to Glutaredoxin
ORF3 Os06g0659800 Conserved hypothetical protein
ORF4 Os06g0660200 Similar to Auxin efflux carrier protein
ORF5 Os06g0660400 Conserved hypothetical protein
ORF6 Os06g0660600 Homeodomain-like containing protein
ORF7 Os06g0660700 Similar to Ubiquitin-conjugating enzyme E2S
ORF8 Os06g0660800 Protein kinase domain containing protein
ORF9 Os06g0661000 Plant MuDR transposase domain containing protein
ORF10 Os06g0661400 ANTH domain containing protein
ORF11 Os06g0661500 Conserved hypothetical protein
ORF12 Os06g0661600 Zinc finger, DHP-type domain containing protein
ORF13 Os06g0661700 RabGAP/TBC domain containing protein
ORF14 Os06g0661800 Similar to Cryptochrome dash
ORF15 Os06g0661900 Protein of unknown function DUF266, plant family protein
ORF16 Os06g0662000 Conserved hypothetical protein
ORF17 Os06g0662200 Eukaryotic transcription factor, DNA-binding domain containing protein
ORF18 Os06g0662300 Pollen allergen Lol p2 family protein
ORF19 Os06g0662500 Pollen allergen Lol p2 family protein
ORF20 Os06g0662600 Pollen allergen Lol p2 family protein
ORF21 Os06g0662700 Pollen allergen Lol p2 family protein
ORF22 Os06g0662800 Pollen allergen Lol p2 family protein
ORF23 Os06g0662900 Pollen allergen/expansion, C-terminal domain containing protein
ORF24 Os06g0663100 Hypothetical protein
ORF25 Os06g0663200 Similar to Protein kinase APK1B, chloroplast precursor
ORF26 Os06g0663300 Pollen allergen Lol p2 family protein
ORF27 Os06g0663400 Serine/threonine protein kinase-like protein

Fig. 6

Expression of WE2 candidate genes"

Fig. 7

Expression analysis of representative genes coding for starch synthesis qRT-PCR assay of the expression of WE2 candidate genes in 12 DAF endosperm. Actin was used as control. n = 3. Values are means ± SD, using Student’s t-test. ** indicates P < 0.01."

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