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Acta Agronomica Sinica ›› 2021, Vol. 47 ›› Issue (5): 860-868.doi: 10.3724/SP.J.1006.2021.04153

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

Genetic and cytological analysis of male sterile mutant msm2015-1 in mungbean

WU Ran-Ran(), LIN Yun, CHEN Jing-Bin, XUE Chen-Chen, YUAN Xing-Xing, YAN Qiang, GAO Ying, LI Ling-Hui, ZHANG Qin-Xue, CHEN Xin*()   

  1. Institute of Industrial Crops, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, Jiangsu, China
  • Received:2020-07-11 Accepted:2020-10-14 Online:2021-05-12 Published:2020-10-30
  • Contact: CHEN Xin E-mail:rrwu@jaas.ac.cn;cx@jaas.ac.cn
  • Supported by:
    National Key Research and Development Program of China(2019YFD1001301);National Key Research and Development Program of China(2019YFD1001300);Natural Science Foundation of Jiangsu Province(BK20190257);National Natural Science Foundation of China(31871696);Jiangsu Agricultural Industry Technology System(JATS[2019]399)

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Abstract:

In China, the low yield per unit has always been a problem in the mungbean industry. The heterosis of mungbean is an effective way to increase yield. The male sterile line is a valuable resource to realize the utilization of hybrid. In this study, 60Co-γ radiation mutagenesis technology was used to induce male sterile mutants in mungbean, and we gained one mutant named male sterile mungbean 2015-1 (msm2015-1) for the first time. There was no significant phenotype difference between msm2015-1 and Sulyu 1 during the vegetative growth stage. The floral organ of msm2015-1 was normal, but the anthers of msm2015-1 were white and could not crack and disperse powder normally in the flowering stage. In the mature stage, hardly any normal pods grew up. Genetic analysis showed the separation ratio of fertility and sterility in the fertile segregation population was in line with 3:1, indicating that the sterility trait was controlled by a single recessive nuclear gene. Cytological analysis of pollen development revealed that there were few pollen grains of msm2015-1 attached to mature stigmas. The pollen grains of msm2015-1 did not germinate in vitro. The cytoplasm of msm2015-1’s pollen could not be stained by Alexander dye. Three abortion types showed up when stained by I2-KI dye: typical abortion, spherical abortion and stained abortion. DAPI staining showed the nuclei of msm2015-1’s pollen developed abnormally. Magenta acetate staining indicated that abortion occurred in the early stage of pollen development, while the asymmetry and more anomalous fission of meiosis during the tetrad stage were the main causes of pollen abortion.

Key words: mungbean, male sterile, genetic analysis, pollen development, cytology

Fig. 1

Phenotypic analysis of the msm2015-1 mutant in mungbean A: vegetative growth stage; B: flower buds; C: opening flowers; D: anatomy of the flower bud, from left to right in order: standard petals, wing petals, keel petals, stamens and pistils; E: the little pods on 2nd day after flowering; F: nearly ripen pods. In figure A, bar = 10 cm; in figure B to F, bar = 1 cm."

Fig. 2

Morphology observation of msm2015-1’s anthers and pollen grains in mungbean A: stamen, the white arrows indicate the indehiscent anther of msm2015-1; B: the pistil of Sulyu 1, the white arrow indicates the attached pollen grains on stigma; C: the pistil of msm2015-1, the white arrow indicates rarely pollen grains attached on stigma; D: scanning electron micrograph (SEM) image of Sulyu 1’s anther; E: SEM image of msm2015-1’s anther; F: SEM image of Sulyu 1’s mature pollen grains; G: SEM image of msm2015-1’s mature pollen grains. In figure A to C, bars = 1 mm; in figure D and E, bars = 100 µm; in fugure F and G, bars = 10 µm."

Table 1

Genetic analysis of fertility separation population"

表型
Phenotype		 观察值
Observed number		 理论值
Theoretical number		 χ2
(3:1)
可育株Fertile plants 187 193.5 0.873
不育株Sterile plants 71 64.5

Fig. 3

Pollen germination of the msm2015-1 mutant in vivo and in vitro in mungbean A: pollen germination on stigma of Sulyu 1, the white arrows indicate the pollen grains attached on stigma and germinated pollen tubes; B: pollen germination on stigma of msm2015-1, the white arrow indicates few pollen grains attached on stigma; C: mature pollen germination of Sulyu 1 in vitro; D: mature pollen germination of msm2015-1 in vitro. Bar = 100 µm."

Fig. 4

Pollen viability of the msm2015-1 mutant analyzed by staining analysis in mungbean A: Alexander staining of Sulyu 1’s mature pollen; B: Alexander staining of msm2015-1’s mature pollen; C: statistics of pollen abortion rate of Alexander staining; D: I2-KI staining of Sulyu 1’s mature pollen; E: I2-KI staining of msm2015-1’s mature pollen; F: statistics of pollen abortion rate of I2-KI staining; G: Alexander staining of Sulyu 1’s anthers of mature flower bud; H: Alexander staining of msm2015-1’s anthers of mature flower bud. Bar = 100 µm."

Fig. 5

Detection of pollen nuclear development in the msm2015-1 mutant A: DAPI staining of Sulyu 1’s mature pollen, the white arrow indicates the generative nucleus, and the red arrow indicates the vegetative nucleus; B: DAPI staining of msm2015-1’s mature pollen. Bar = 20 µm."

Fig. 6

Developmental early stage analysis of msm2015-1’s pollen in mungbean A: the early stages of pollen development, the white arrows indicate the uneven-size microspores caused by the asymmetry and inhomogeneity meiosis during the tetrad stage; B: the anomalous more fission of meiosis happened during the tetrad stage, the numbers of msm2015-1’s microspores are indicated by the Arabic numbers. PMC: pollen mother cell; Msp: microspore. Bar = 20 µm."

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