Welcome to Acta Agronomica Sinica，

Acta Agronomica Sinica ›› 2021, Vol. 47 ›› Issue (5): 983-990.doi: 10.3724/SP.J.1006.2021.04207

• RESEARCH NOTES • Previous Articles    

Morphological characteristics and cytological study of anther abortion of temperature-sensitive nuclear male sterile line 160S in Brassica napus

TANG Xin(), LI Yuan-Yuan, LU Jun-Xing, ZHANG Tao*()   

  1. College of Life Sciences, Chongqing Normal University, Chongqing 401331, China
  • Received:2020-09-09 Accepted:2020-11-13 Online:2021-05-12 Published:2020-12-15
  • Contact: ZHANG Tao E-mail:1047910732@qq.com;zht2188@126.com
  • Supported by:
    Natural Science Foundation of Chongqing(cstc2012jjA80010);Science and Technology Bureau Foundation Project Chongqing(cstc2017jcyjAX0108);Scientific and Technological Research Program of Chongqing Municipal Education Commission(KJQN201900533)

RichHTML382

23

PDF

550

Abstract:

The main purpose of this study was to explore the flower morphologies, anther abortion times and cytological characteristics of the thermo sensitive genic male sterile line 160S in Brassica napus. The results will provide a theoretical basis for further study on the molecular regulation mechanism and guide significance for its practical application in two-line hybrid breeding of rape. The 160S were grown at 15℃ and 28℃. The flower morphology, microspore, anther structure and tapetum at different stages of fertile plants (Male Fertile/160S-MF) and sterile plants (Male Sterile/160S-MS) were observed by stereomicroscope with carmine acetate staining, paraffin section, hematoxylin eosin staining and TUNEL staining. The anther was yellow, plump and dehiscent with high pollen viability at 15℃. At 28℃, there was no differences in pistil and potato chips between 160S-MS and 160S-MF flowers, but the petals and the filaments of 160S-MS were smaller and shorter. The stamens of 160S-MS degenerated obviously and its anthers were shriveled in yellowish brown without pollen grains, indicating completely male sterility. But interestingly, the 160S-MF developed normally with higher pollen viability. However, microspore and pollen grains were not observed in 160S-MS due to complete abortion of stamens. Paraffin section results showed that there was no significant difference between 160S-MS and 160S-MF in sporulation and pollen mother cell stages. The morphology and structure of tapetum of 160S-MS appeared some abnormal appearances, such as irregularly cell arrangement, vacuolization, and premature disintegration at meiosis stage. Meanwhile, the development of pollen mother cells were hindered and no tetrads structure were formed leading to the empty pollen sac before the completion of meiosis. TUNEL assay showed that the tapetum of 160-MS began to apoptosis at meiosis stage. In summary, these results showed that 160S belongs to pollen mother cell abortion type male sterile line, whose abortion stage occured during meiosis stage and the tapetum cells degraded prematurely and failed to be transformed to the glandular type, so that the nutrients needed for pollen mother cell development were not provided. The tetrad structures and microspores can not be formed, which leaded to the formation of empty anther sac and resulted in male sterility.

Key words: Brassica napus, thermo-sensitive genic male sterility, anther abortion, cytological study

Fig. 1

Flower morphology of 160S-MF and 160S-MS A: the flower of 160S-MF; B: the flower stamens of 160S-MF; C: the petal and sepal of 160S-MF; D: the flower of 160S-MS; E: the flower stamens of 160S-MS; F: the petal and sepal of 160S-MS."

Table 1

Comparison of the flower traits between 160S-MF and 160S-MS (mm)"

花器特征
Flower trait		 育性类型Types of male sterility
160S-MF 160S-MS
花冠径Diameter of corolla 18.97±1.68 15.58±0.78**
花瓣长Length of petal 13.98±1.16 12.30±0.57**
花瓣宽Width of petal 7.48±0.78 5.88±0.37**
雌蕊长Length of pistil 9.24±1.13 9.80±0.72
四强雄蕊长Length of four stronger stamens 9.38±0.45 6.11±1.09**
二弱雄蕊长Length of two weaker stamens 6.99±0.81 4.89±1.20**
四强花药长Length of four stronger stamens’ anther 2.25±0.15 1.80±0.08**
花丝长Length of four long filaments 7.44±0.46 4.69±0.89**
花柄长Length of stalk 29.13±1.38 16.50±1.27**
萼片长Length of sepal 7.10±0.82 6.61±0.51

Fig. 2

Microspores morphology of 160S-MF and 160S-MS A: terads of 160S-MF; B: early microspores of 160S-MF; C: uninucleate microspores of 160S-MF; D: binucleate microspores of 160S-MF; E: pollen grains of 160S-MF; F: pollen grains of 160S-MS. The arrow points to the nucleus of the cells."

Fig. 3

Microscopical structure of anthers at different developmental stages in 160S-MF A, B: sporogenous cell stage; C: pollen mother cell stage; D: meiotic stage; E: tetrad stage; F: late uninucleate stage; G: binucleate stage; H, I: mature pollen; Epi: epidermal; Mid: middle layer; En: endothecium; T: tapetum; Sp: sporogenous cell; PMC: pollen mother cell; Te: tetrad; M: microspore; Po: pollen."

Fig. 4

Microscopical structure of anthers at different developmental stages in 160S-MS A, B: sporogenous cell stage; C: pollen mother cell stage; D, E: meiotic stage; F: locule of aborted anther; Epi: epidermal; Mid: middle layer; En: endothecium; T: tapetum; Sp: sporogenous cell; PMC: pollen mother cell."

Fig. 5

TUNEL detection of anther programmed cell dead at different developmental stages in 160S-MF and 160S-MS A: 160S-MF at pollen mother cell stage; B: 160S-MS at pollen mother cell stage; C: 160S-MF at meiotic stage; D and E:160S-MS at meiotic stage; F: 160S-MF at tetrad stage; G:160S-MF at binucleate stage; T: tapetum; PMC: pollen mother cell; Te: tetrad; M: microspore. The fluoresce of nuclei stained by propidium iodide is red, while green fluorescence is TUNEL-positive nuclei staining (arrow)."

[1] Tester M, Langridge P. Breeding technologies to increase crop production in a changing world. Science, 2010,327:818-822.
pmid: 20150489
[2] Rondanini D P, Gomez N V, Agosti M B, Miralles D J. Global trends of rapeseed grain yield stability and rapeseed-to-wheat yield ratio in the last four decades. Eur J Agron, 2012,37:56-65.
[3] Hochholdinger F, Hoecker N. Towards the molecular basis of heterosis. Trends Plant Sci, 2007,12:427-432.
doi: 10.1016/j.tplants.2007.08.005 pmid: 17720610
[4] Chen L, Liu Y G. Male sterility and fertility restoration in crops. Ann Rev Plant Biol, 2014,65:579-606.
[5] Virmani S S, Ilyas-Ahmed M. Environment-sensitive genic male sterility (EGMS) in crops. Adv Agron, 2001,72:139-195.
[6] Lei S L, Yao X Q, Yi B, Chen W, Ma C Z, Tu J X, Fu T D. Towards map-based cloning: fine mapping of a recessive genic male-sterile gene ( BnMs2) in Brassica napus L. and syntenic region identification based on the Arabidopsis thaliana genome sequences. Theor Appl Genet, 2007,115:643-651.
doi: 10.1007/s00122-007-0594-1 pmid: 17605126
[7] Sun Y, Zhang D, Wang Z, Guo Y, Sun X, Li W, Zhi W, Hu S. Cytological observation of anther structure and genetic investigation of a thermo-sensitive genic male sterile line 373S in Brassica napus L. BMC Plant Biol, 2020,20:8.
[8] 石明松. 对光照长度敏感的隐性雄性不育水稻的发现与初步研究. 中国农业科学, 1985,18(2):44-48.
Shi M S. The discovery and study of the photosensitive recessive male-sterile rice ( Oryza sativa L. subsp. japonica). Sci Agric Sin, 1985,18(2):44-48 (in Chinese with abstract).
[9] Lee D S, Chen L J, Suh H S. Genetic characterization and fine mapping of a novel thermo-sensitive genic male-sterile gene tms6 in rice ( Oryza sativa L.). Theo Appl Genet, 2005,111:1271-1277.
[10] Tang J H, Fu Z Y, Hu Y M, Li J S, Sun L L, Ji H Q. Genetic analyses and mapping of a new thermo-sensitive genic male sterile gene in maize. Theor Appl Genet, 2006,113:11-15.
doi: 10.1007/s00122-006-0262-x pmid: 16783588
[11] Xing Q H, Ru Z G, Zhou C J, Xue X, Liang C Y, Yang D E, Jin D M, Wang B. Genetic analysis, molecular tagging and mapping of the thermo-sensitive genic male-sterile gene ( wtms1) in wheat. Theor Appl Genet, 2003,107:1500-1504.
doi: 10.1007/s00122-003-1385-y pmid: 12928780
[12] Frasch R M, Weigand C, Perez P T, Palmer R G, Sandhu D. Molecular mapping of 2 environmentally sensitive male-sterile mutants in soybean. J Hered, 2011,102:11-16.
doi: 10.1093/jhered/esq100 pmid: 20864624
[13] Zhu J, Lou Y, Shi Q S, Zhang S, Zhou W T, Yang J, Zhang C, Yao X Z, Xu T, Liu J L, Zhou L, Hou J Q, Wang J Q, Wang S, Huang X H, Yang Z N. Slowing development restores the fertility of thermo-sensitive male-sterile plant lines. Nat Plants, 2020,6:360-367.
pmid: 32231254
[14] 李可琪, 曾新华, 袁荣, 闫晓红, 吴刚. 甘蓝型油菜温敏细胞核雄性不育系TE5A花药发育的细胞学研究. 中国农业科学, 2016,49:2408-2417.
Li K Q, Zeng X H, Yuan R, Yan X H, Wu G. Cytological researches on the anther development of a thermo-sensitive genic male sterile line TE5A in Brassica napus. Sci Agric Sin, 2016,49:2408-2417 (in Chinese with English abstract).
[15] Yan X, Zeng X, Wang S, Li K, Yuan R, Gao H, Luo J, Liu F, Wu Y, Li Y, Zhu L, Wu G. Aberrant meiotic prophase I leads to genic male sterility in the novel TE5A mutant of Brassica napus. Sci Rep, 2016,6:33955.
pmid: 27670217
[16] Yu C Y, Guo Y F, Ge J, Hu Y M, Dong J G, Dong Z S. Characterization of a new temperature-sensitive male sterile line SP2S in rapeseed ( Brassica napus L.). Euphytica, 2015,206:473-485.
[17] 张涛, 沈余亮, 王瑞雪, 邹燕, 赵敬会, 李荣冲, 梁晶龙, 龚慧明. 甘蓝型油菜雄性不育系160S育性转换与利用. 西北植物学报, 2012,32:35-41.
Zhang T, Shen L Y, Wang R X, Zou Y, Zhao J H, Li R C, Liang J L, Gong H M. Fertility alteration and utilization of male-sterile line 160S in Brassica napus. Acta Bot Boreali-Occident Sin, 2012,32:35-41 (in Chinese with English abstract).
[18] Laser K D, Lersten N K. Anatomy and cytology of microsporo-genesis in cytoplasmic male sterile (CMS) angiosperms. Bot Rev, 1972,38:425-454.
[19] Wang S P, Zhang G S, Song Q L, Zhang Y X, Li Z, Guo J L, Niu N, Ma S C, Wang J W. Abnormal development of tapetum and microspores induced by chemical hybridization agent SQ-1 in wheat. PLoS One, 2015,10:e0119557.
doi: 10.1371/journal.pone.0119557 pmid: 25803723
[20] 余凤群, 傅廷栋. 甘蓝型油菜几个雄性不育系花药发育的细胞形态学研究. 武汉植物学研究, 1990,8:209-216.
Yu F Q, Fu T D. Cytomorphology study on anther development of several male sterile lines of Brassica napus. J Wuhan Bot Res, 1990,8:209-216 (in Chinese with English abstract).
[21] 葛娟, 郭英芬, 于澄宇, 张国云, 董军刚, 董振生. 甘蓝型油菜光、温敏雄性不育系Huiyou 50S花粉败育的细胞学观察. 作物学报, 2012,38:541-548.
Ge J, Guo Y F, Yu C Y, Zhang G Y, Dong J G, Dong Z S. Cytological observation of anther development of photoperiod/thermo-sensitive male sterile line Huiyou 50S in Brassica napus. Acta Agron Sin, 2012,38:541-548 (in Chinese with English abstract).
[22] Ma H. Molecular genetic analyses of microsporogenesis and microgametogenesis in flowering plants. Annu Rev Plant Biol, 2005,56:393-434.
pmid: 15862102
[23] Jung K H, Han M J, Lee Y S, Kim Y W, Hwang I W, Kim M J, Kim Y K, Nahm B H, An G. Rice undeveloped tapetum1 is a major regulator of early tapetum development. Plant Cell, 2005,17:2705-2722.
doi: 10.1105/tpc.105.034090 pmid: 16141453
[24] Zhao D Z, Wang G F, Speal B, Ma H. The excess microsporocytes1 gene encodes a putative leucine-rich repeat receptor protein kinase that controls somatic and reproductive cell fates in the Arabidopsis anther. Genes Dev, 2002,16:2021-2031.
pmid: 12154130
[25] Bedinger P. The remarkable biology of pollen. Plant Cell, 1992,4:879-887.
[26] Xu J, Ding Z W, Vizcay-Barrena G, Shi J X, Liang W Q, Yuan Z, Werck-Reichhart D, Schreiber L, Wilson Z A, Zhang D B. Aborted microspores acts as a master regulator of pollen wall formation in Arabidopsis. Plant Cell, 2014,26:1544-1556.
doi: 10.1105/tpc.114.122986 pmid: 24781116
[27] Liu Z H, Shi X Y, Li S, Hu G, Zhang L L, Song X Y. Tapetal-delayed programmed cell death (PCD) and oxidative stress-induced male sterility of Aegilops uniaristata cytoplasm in wheat. Int J Mol Sci, 2018,19:1708.
[28] Papini A, Mosti S, Brighigna L. Programmed-cell-death events during tapetum development of angiosperms. Protoplasma, 1999,207:213-221.
[29] Li N, Zhang D S, Liu H S, Yin C S, Li X X, Liang W Q, Yuan Z, Xu B, Chu H W, Wang J, Wen T Q, Huang H, Luo D, Ma H, Zhang D B. The ricetapetum degeneration retardation gene is required for tapetum degradation and anther development. Plant Cell, 2006,18:2999-3014.
[30] Zhang D, Liu D, Lyu X, Wang Y, Xun Z, Liu Z, Li F, Lu H. The cysteine protease CEP1, a key executor involved in tapetal programmed cell death, regulates pollen development in Arabidopsis. Plant Cell, 2014,26:2939-2961.
[31] Jacobowitz J R, Doyle W C, Weng J K. PRX9 and PRX40 are extensin peroxidases essential for maintaining tapetum and microspore cell wall integrity during Arabidopsis anther development. Plant Cell, 2019,31:848-861.
doi: 10.1105/tpc.18.00907 pmid: 30886127
[32] Xie X, Zhang Z, Zhao Z, Xie Y, Li H, Ma X, Liu Y G, Chen L. The mitochondrial aldehyde dehydrogenase OsALDH2b negatively regulates tapetum degeneration in rice. J Exp Bot, 2020,71:2551-2560.
doi: 10.1093/jxb/eraa045 pmid: 31989154
[33] Uzair M, Xu D, Schreiber L, Shi J, Liang W, Jung K H, Chen M, Luo Z, Zhang Y, Yu J, Zhang D. PERSISTENT TAPETAL CELL2 is required for normal tapetal programmed cell death and pollen wall patterning. Plant Physiol, 2020,182:962-976.
pmid: 31772077
[34] 许代香, 贾乐东, 王瑞, 马国强, 段谋正, 曲存民, 李加纳. 甘蓝型油菜显性核不育系D3A的细胞学研究. 西南大学学报(自然科学版), 2020,42(1):16-21.
Xu D X, Jia L D, Wang R, Ma G Q, Duan M Z, Qu C M, Li J N. Cytological studies of dominant GMS sterile line D3A in Brassica napus L. J Southwest Univ (Nat Sci Edn), 2020,42(1):16-21 (in Chinese with English abstract).
[35] 李六林. 植物雄性不育机理研究. 北京: 中国农业科学技术出版社, 2008. p 5.
Li L L. Study on the Mechanism of Plant Male Sterility. Beijing: China Agricultural Science and Technology Press, 2008. p 5 (in Chinese).
[1] CHEN Song-Yu, DING Yi-Juan, SUN Jun-Ming, HUANG Deng-Wen, YANG Nan, DAI Yu-Han, WAN Hua-Fang, QIAN Wei. Genome-wide identification of BnCNGC and the gene expression analysis in Brassica napus challenged with Sclerotinia sclerotiorum and PEG-simulated drought [J]. Acta Agronomica Sinica, 2022, 48(6): 1357-1371.
[2] YUAN Da-Shuang, DENG Wan-Yu, WANG Zhen, PENG Qian, ZHANG Xiao-Li, YAO Meng-Nan, MIAO Wen-Jie, ZHU Dong-Ming, LI Jia-Na, LIANG Ying. Cloning and functional analysis of BnMAPK2 gene in Brassica napus [J]. Acta Agronomica Sinica, 2022, 48(4): 840-850.
[3] HUANG Cheng, LIANG Xiao-Mei, DAI Cheng, WEN Jing, YI Bin, TU Jin-Xing, SHEN Jin-Xiong, FU Ting-Dong, MA Chao-Zhi. Genome wide analysis of BnAPs gene family in Brassica napus [J]. Acta Agronomica Sinica, 2022, 48(3): 597-607.
[4] WANG Rui, CHEN Xue, GUO Qing-Qing, ZHOU Rong, CHEN Lei, LI Jia-Na. Development of linkage InDel markers of the white petal gene based on whole-genome re-sequencing data in Brassica napus L. [J]. Acta Agronomica Sinica, 2022, 48(3): 759-769.
[5] WANG Yan-Hua, LIU Jing-Sen, LI Jia-Na. Integrating GWAS and WGCNA to screen and identify candidate genes for biological yield in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(8): 1491-1510.
[6] LI Jie-Hua, DUAN Qun, SHI Ming-Tao, WU Lu-Mei, LIU Han, LIN Yong-Jun, WU Gao-Bing, FAN Chu-Chuan, ZHOU Yong-Ming. Development and identification of transgenic rapeseed with a novel gene for glyphosate resistance [J]. Acta Agronomica Sinica, 2021, 47(5): 789-798.
[7] ZHOU Xin-Tong, GUO Qing-Qing, CHEN Xue, LI Jia-Na, WANG Rui. Construction of a high-density genetic map using genotyping by sequencing (GBS) for quantitative trait loci (QTL) analysis of pink petal trait in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(4): 587-598.
[8] LI Shu-Yu, HUANG Yang, XIONG Jie, DING Ge, CHEN Lun-Lin, SONG Lai-Qiang. QTL mapping and candidate genes screening of earliness traits in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(4): 626-637.
[9] TANG Jing-Quan, WANG Nan, GAO Jie, LIU Ting-Ting, WEN Jing, YI Bin, TU Jin-Xing, FU Ting-Dong, SHEN Jin-Xiong. Bioinformatics analysis of SnRK gene family and its relation with seed oil content of Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(3): 416-426.
[10] MENG Jiang-Yu, LIANG Guang-Wei, HE Ya-Jun, QIAN Wei. QTL mapping of salt and drought tolerance related traits in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(3): 462-471.
[11] WEI Li-Juan, SHEN Shu-Lin, HUANG Xiao-Hu, MA Guo-Qiang, WANG Xi-Tong, YANG Yi-Ling, LI Huan-Dong, WANG Shu-Xian, ZHU Mei-Chen, TANG Zhang-Lin, LU Kun, LI Jia-Na, QU Cun-Min. Genome-wide association analysis reveals zinc-tolerant loci of rapeseed at germination stage [J]. Acta Agronomica Sinica, 2021, 47(2): 262-274.
[12] LI Qian, Nadil Shah, ZHOU Yuan-Wei, HOU Zhao-Ke, GONG Jian-Fang, LIU Jue, SHANG Zheng-Wei, ZHANG Lei, ZHAN Zong-Xiang, CHANG Hai-Bin, FU Ting-Dong, PIAO Zhong-Yun, ZHANG Chun-Yu. Breeding of a novel clubroot disease-resistant Brassica napus variety Huayouza 62R [J]. Acta Agronomica Sinica, 2021, 47(2): 210-223.
[13] WANG Rui-Li, WANG Liu-Yan, LEI Wei, WU Jia-Yi, SHI Hong-Song, LI Chen-Yang, TANG Zhang-Lin, LI Jia-Na, ZHOU Qing-Yuan, CUI Cui. Screening candidate genes related to aluminum toxicity stress at germination stage via RNA-seq and QTL mapping in Brassica napus L. [J]. Acta Agronomica Sinica, 2021, 47(12): 2407-2422.
[14] WANG Zhen, ZHANG Xiao-Li, MENG Xiao-Jing, YAO Meng-Nan, MIU Wen-Jie, YUAN Da-Shuang, ZHU Dong-Ming, QU Cun-Min, LU Kun, LI Jia-Na, LIANG Ying. Identification of upstream regulators for mitogen-activated protein kinase 7 gene (BnMAPK7) in rapeseed (Brassica napus L.) [J]. Acta Agronomica Sinica, 2021, 47(12): 2379-2393.
[15] GUO Qing-Qing, ZHOU Rong, CHEN Xue, CHEN Lei, LI Jia-Na, WANG Rui. Location and InDel markers for candidate interval of the orange petal gene in Brassica napus L. by next generation sequencing [J]. Acta Agronomica Sinica, 2021, 47(11): 2163-2172.
Viewed
Full text


Abstract

Cited
  Shared   
  Discussed   
No Suggested Reading articles found!
Add: No. 80 Xueyuan South Rd, Beijing 100081, P. R. China E-mail: zwxb301@caas.cn
Supported by Beijing Magtech Co.Ltd