Optimization of Seed Sterilization and Rooting Medium for Regeneration of <i>Brassica napus</i>

FU Shu; LIU Zhao-xia; CHEN Jin-zhi; SUN Geng-xiao; SUN Cui-ying; CHEN Jin; YANG Guang

doi:10.19303/j.issn.1008-0384.2019.12.003

Volume 34 Issue 12

Dec. 2019

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Article Navigation > Fujian Journal of Agricultural Sciences > 2019 > 34(12): 1371-1378

FU S, LIU Z X, CHEN J Z, et al. Optimization of Seed Sterilization and Rooting Medium for Regeneration of Brassica napus [J]. Fujian Journal of Agricultural Sciences，2019，34（12）：1371−1378. doi: 10.19303/j.issn.1008-0384.2019.12.003

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FU S, LIU Z X, CHEN J Z, et al. Optimization of Seed Sterilization and Rooting Medium for Regeneration of Brassica napus [J]. Fujian Journal of Agricultural Sciences，2019，34（12）：1371−1378. doi: 10.19303/j.issn.1008-0384.2019.12.003

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Optimization of Seed Sterilization and Rooting Medium for Regeneration of Brassica napus

doi: 10.19303/j.issn.1008-0384.2019.12.003

FU Shu^{1, 2, 3
,},
LIU Zhao-xia^{1, 2, 3},
CHEN Jin-zhi^{1, 2, 3},
SUN Geng-xiao^{1, 2, 3},
SUN Cui-ying^{1, 2, 3},
CHEN Jin^{1, 2, 3},
YANG Guang^{1, 2, 3
,
,}

1.
State Key Laboratory of Ecological Pest Control for Fujian and Taiwan Crops/ Institute of Applied Ecology, Fuzhou, Fujian　350002, China
2.
Joint International Research Laboratory of Ecological Pest Control, Ministry of Education, Fuzhou, Fujian　350002, China
3.
Key Laboratory of Green Control of Insect Pests (Fujian Province University), Fuzhou, Fujian　350002, China

Received Date: 2019-11-18
Rev Recd Date: 2019-12-09
Publish Date: 2019-12-01

Abstract

Abstract

Objective Seed sterilization and rooting culture are important steps in plant tissue culture. At present, there is considerable variation in the concentration of sodium hypochlorite (NaClO) and seed sterilization time of Brassica napus, as well as the type and concentration of hormone in the rooting media. The purpose of this study was to optimize the method of seed sterilization with NaClO and the formula of rooting medium for B. napus regeneration. Method The optimum concentration of NaClO for seed sterilization was screened by comparing the growth status (such as germination rate, healthy shoot rate and rate of colonies) of seeds treated with different concentrations (0.1%–30.0%) of NaClO, and the appropriate sterilization time was optimized based on the growth status of seeds treated with the optimal concentration of NaClO for 10—30 min. Moreover, the optimum formula of rooting media was screened by comparing the rooting regeneration of seedlings in the media containing 0.1–2.0 mg·L⁻¹ naphthylacetic acid (NAA) / indolebutyric acid (IBA). Result The optimum concentration of NaClO for seed sterilization was 2.0% or 3.0%, and the best time for seed sterilization with 3.0% NaClO was 15 min, causing the 97.4% of germination rate and 85.9% of healthy shoot rate with least rate of colonies. Moreover, this method of seed sterilization is applicable to other 9 plants, especially for B. napus, B. pekinensis and Arabidopsis thaliana. The root regenerated best in the rooting medium containing 0.1 mg·L⁻¹ NAA, in which 92.5% of root regenerated from the cutting point on the 7^th day with the most lateral roots. Conclusion This study optimized the method of seed sterilization and the formula of rooting medium, which laid a foundation for efficient plant tissue culture of B. napus.
- Brassica napus,
- seed sterilization,
- sodium hypochlorite,
- rooting,
- NAA,
- IBA

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References(32)

References

[1]	OYEBANJI O, NWEKE O, ODEBUNMI O, et al. Simple, effective and economical explant-surface sterilization protocol for cowpea, rice and sorghum seeds [J]. African Journal of Biotechnology, 2009, 8(20): 5395−5399.
[2]	YILDIZ M, EKIZ H. The effect of sodium hypochlorite solutions on in vitro seedling growth and regeneration capacity of sainfoin (Onobrychis viciifolia Scop.) hypocotyl explants [J]. Canadian Journal of Plant Science, 2014, 94(7): 1161−1164. doi: 10.4141/cjps2013-250
[3]	郝梦宇, 丁炳莉, 李超, 等. 甘蓝型油菜组培苗生根培养体系的优化 [J]. 中国油料作物学报, 2018, 40(3):352−358. doi: 10.7505/j.issn.1007-9084.2018.03.006 HAO M Y, DING B L, LI C, et al. Optimization of rooting medium for in vitro transgenic shoots in Brassica napus [J]. Chinese Journal of Oil Crop Sciences, 2018, 40(3): 352−358.(in Chinese) doi: 10.7505/j.issn.1007-9084.2018.03.006
[4]	ZIAEI M, MOTALLEBI M, ZAMANI M R, et al. Co-expression of chimeric chitinase and a polygalacturonase-inhibiting protein in transgenic canola (Brassica napus) confers enhanced resistance to Sclerotinia sclerotiorum [J]. Biotechnology Letters, 2016, 38(6): 1021−1032. doi: 10.1007/s10529-016-2058-7
[5]	BHALLA P L, SINGH M B. Agrobacterium-mediated transformation of Brassica napus and Brassica oleracea [J]. Nature Protocols, 2008, 3(2): 181−189. doi: 10.1038/nprot.2007.527
[6]	RADKE S E, ANDREWS B M, MOLONEY M M, et al. Transformation of Brassica napus L. using Agrobacterium tumefaciens: developmentally regulated expression of a reintroduced napin gene [J]. Theoretical and Applied Genetics, 1988, 75(5): 685−694. doi: 10.1007/BF00265588
[7]	FAN Y, DU K, GAO Y, et al. Transformation of LTP gene into Brassica napus to enhance its resistance to Sclerotinia sclerotiorum [J]. Russian Journal of Genetics, 2013, 49(4): 380−387. doi: 10.1134/S1022795413040042
[8]	SONNTAG K. Genotype and procedure dependence of Agrobacterium-mediated transformation of Brassica napus [J]. Journal of Consumer Protection and Food Safety, 2007, 2(1): 113.
[9]	ENGELKE T, HIRSCHE J, ROITSCH T. Metabolically engineered male sterility in rapeseed (Brassica napus L.) [J]. Theoretical and Applied Genetics, 2011, 122(1): 163−174. doi: 10.1007/s00122-010-1432-4
[10]	KONG F, LI J, TAN X, et al. New time-saving transformation system for Brassica napus [J]. African Journal of Biotechnology, 2009, 8(11): 2497−2502.
[11]	WANG Y Q, ZHANG Y, WANG F, et al. Development of transgenic Brassica napus with an optimized cry1C* gene for resistance to diamondback moth (Plutella xylostella) [J]. Canadian Journal of Plant Science, 2014, 94(8): 1501−1506. doi: 10.4141/cjps-2014-099
[12]	LIU H B, GUO X, NAEEM M S, et al. Transgenic Brassica napus L. lines carrying a two gene construct demonstrate enhanced resistance against Plutella xylostella and Sclerotinia sclerotiorum [J]. Plant Cell, Tissue and Organ Culture, 2011, 106(1): 143−151. doi: 10.1007/s11240-010-9902-6
[13]	ELHITI M, YANG C, CHAN A, et al. Altered seed oil and glucosinolate levels in transgenic plants overexpressing the Brassica napus SHOOTMERISTMLESS gene [J]. Journal of Experimental Botany, 2012, 63(12): 4447−4461. doi: 10.1093/jxb/ers125
[14]	MCALLISTER C H, WOLANSKY M, GOOD A G. The impact on nitrogen-efficient phenotypes when aspartate aminotransferase is expressed tissue-specifically in Brassica napus [J]. New Negatives in Plant Science, 2016, 3: 1−9.
[15]	WANG Y, XU H, KOU J J, et al. Dual effects of transgenic Brassica napus overexpressing CS gene on tolerances to aluminum toxicity and phosphorus deficiency [J]. Plant Soil, 2013, 362: 231−246. doi: 10.1007/s11104-012-1289-1
[16]	JIANG Y Z, FU X L, WEN M L, et al. Overexpression of an nsLTPs-like antimicrobial protein gene (LJAMP2) from motherwort (Leonurus japonicus) enhances resistance to Sclerotinia sclerotiorum in oilseed rape (Brassica napus) [J]. Physiological and Molecular Plant Pathology, 2013, 82: 81−87. doi: 10.1016/j.pmpp.2012.11.001
[17]	KHAN M R, RASHID H, ANSAR M, et al. High frequency shoot regeneration and Agrobacterium-mediated DNA transfer in Canola (Brassica napus) [J]. Plant Cell, Tissue and Organ Culture, 2003, 75(3): 223−231. doi: 10.1023/A:1025869809904
[18]	MASHAYEKHI M, SHAKIB A M, AHMADRAJI M, et al. Gene transformation potential of commercial canola (Brassica napus L.) cultivars using cotyledon and hypocotyl explants [J]. African Journal of Biotechnology, 2008, 7(24): 4459−4463.
[19]	ORLIKOWSKA T, NOWAK K, REED B. Bacteria in the plant tissue culture environment [J]. Plant Cell, Tissue and Organ Culture, 2017, 128(3): 487−508. doi: 10.1007/s11240-016-1144-9
[20]	杜燕, 蒋海玉, 刘其宁. 贮存过期油菜种子消毒方法的研究 [J]. 种子, 2003(2):39−40. doi: 10.3969/j.issn.1001-4705.2003.02.016 DU Y, JIANG H Y, LIU Q L. Studies on rescue and propagation of rape germplasm resources exceeding the effective storage duration the collection of sterilization methods [J]. Seed, 2003(2): 39−40.(in Chinese) doi: 10.3969/j.issn.1001-4705.2003.02.016
[21]	耿思宇, 张姗姗, 徐培林, 等. 激素组合等在甘蓝型油菜下胚轴再生中的作用 [J]. 山西农业科学, 2019, 47(5):730−733, 760. doi: 10.3969/j.issn.1002-2481.2019.05.07 GENG S Y, ZHANG S S, XU P L, et al. Role of hormone combinations on hypocotyls regeneration in Brassica napus [J]. Journal of Shanxi Agricultural Sciences, 2019, 47(5): 730−733, 760.(in Chinese) doi: 10.3969/j.issn.1002-2481.2019.05.07
[22]	杨长友, 袁中厚, 郑小敏, 等. 甘蓝型油菜高效离体再生体系的建立 [J]. 生物技术通报, 2013(1):111−115. YANG C Y, YUAN Z H, ZHENG X M, et al. Establishment of effective regeneration system of Brassica napus L. in vivo [J]. Biotechnology Bulletin, 2013(1): 111−115.(in Chinese)
[23]	黄昌蓉. 甘蓝型油菜早熟基因遗传转化体系的建立及转基因植株初步鉴定[D]. 杭州: 浙江大学, 2013. HUANG C R. Establishment of genetic transformation with earliness gene and preliminary identification of transgenic plant in Brassica napus[D]. Hangzhou: Zhejiang University, 2013. (in Chinese)
[24]	BIJANZADEH E, NOSRATI K, EGAN T. Influence of seed priming techniques on germination and emergence of rapeseed (Brassica napus L.) [J]. Seed Science and Technology, 2010, 38: 242−247. doi: 10.15258/sst.2010.38.1.26
[25]	HATZIG S V, FRISCH M, BREUER F, et al. Genome-wide association mapping unravels the genetic control of seed germination and vigor in Brassica napus [J]. Frontiers in Plant Science, 2015(6): 221.
[26]	LINDSEY B, RIVERO L, CALHOUN C S, et al. Standardized method for high-throughput sterilization of Arabidopsis seeds [J]. Journal of Visualized Experiments, 2017, 128: e56587.
[27]	JOVIČIČ D, POPOVIČ B M, JEROMELA A M, et al. The interaction between salinity stress and seed ageing during germination of Brassica napus seeds [J]. Seed Science and Technology, 2019, 47(1): 47−52. doi: 10.15258/sst.2019.47.1.05
[28]	HAO Z P, HUANG F, HOU S M, et al. Varietal differences in response to imidacloprid seed treatment in germination and early seedling growth of oilseed rape [J]. Seed Science and Technology, 2019, 47(1): 1−12. doi: 10.15258/sst.2019.47.1.01
[29]	MALEK M, GHADERI-FAR F, TORABI B, et al. The influence of seed priming on storability of rapeseed (Brassica napus) seeds [J]. Seed Science and Technology, 2019, 47(1): 87−92. doi: 10.15258/sst.2019.47.1.09
[30]	MAGRINI S, VITIS M D. In vitro reproduction of three Limodorum species (Orchidaceae): impacts of scarification methods and nitrogen sources on mature seed germination and seedling development [J]. Plant Biosystems, 2017, 151(3): 419−428. doi: 10.1080/11263504.2016.1179698
[31]	YADAV K, SINGH N. Effect of seed harvesting season and sterilization treatments on germination and in vitro propagation of Albizia lebbeck (L.) Benth [J]. Analele Universitatii din Oradea, Fascicula Biologie, 2011, 18(2): 151−156.
[32]	DING H, FU T J, SMITH M A. Microbial contamination in sprouts: how effective is seed disinfection treatment? [J]. Journal of Food Science, 2013, 78(4): 495−501. doi: 10.1111/1750-3841.12064