A New Method for Onion Seed Disinfection and Aseptic Seedling Culture

doi:10.13560/j.cnki.biotech.bull.1985.2022-0983

Abstract

Abstract:

This work aims to obtain a simple, efficient and low-contamination method for onion seed disinfection and aseptic seedling culture. The sterilization of onion seeds and the culture of sterile seedlings were studied by controlling the pH value, disinfection soaking time and culture concentration of plant preservative mixture（PPM）. The results showed that the disinfection effect of soaking seeds in PPM（20 mL/L, pH=2）for 24-48 h reached the level of 10% NaClO disinfection for 10 min, which did not affect seed germination, and the germination potential exceeded 90% in 8 d. The analysis of contaminated microorganisms found that four species of fungi（Alternaria alternata, Fusarium proliferatum, Fusarium oxysporum, and Stemphylium vesicarium）and two species of bacteria（Phytobacter diazotrophicus and Atlantibacter hermannii）were transmitted through onion seeds, and the bacteriostasis test confirmed that PPM was able to effectively inhibit the growth of contaminated microorganisms. The medium added with PPM effectively reduced the contamination rate using PPM for seed soaking disinfection; high concentration PPM（1-9 mL/L）inhibited the growth of onion seedlings, while low concentration PPM（0-0.5 mL/L）had no significant inhibitory effect on the growth of onion seedlings. In sum, a set of simple and efficent methods for onion seed disinfection and aseptic seedling culture are explored and developed.

Key words: onion（Allium cepa）, seed, disinfection, aseptic seedling, plant preservative mixture（PPM）

SUN Ya-ling, LI Rui-ping, WANG Zhen-bao, ZHANG Shu, LIU Bing-jiang, HUO Yu-meng. A New Method for Onion Seed Disinfection and Aseptic Seedling Culture[J]. Biotechnology Bulletin, 2023, 39(4): 212-220.

Figures/Tables 9

Fig. 1 Effects of pH value of PPM disinfectant on onion seed germination（A）and disinfection（B） Different letters at the top of the column chart indicate significant differences among different treatments（P<0.05）. The Chi square value and the adjusted P value are marked above the horizontal line. The same below

Fig. 2 Effects of PPM disinfection time on onion seed germination A1：Directly sowed dry seeds to 1/2 MS（1 mL/L PPM）. A2：Sowed seeds to 1/2 MS（1 mL/L PPM）after surface disinfection by 20 mL/L PPM. A3-A7：Sowed seeds to 1/2 MS（1 mL/L PPM）after soaked and disinfected for 3, 6, 12, 24 and 48 h respectively by 20 mL/L PPM. A8：Sowed seeds to 1/2 MS after disinfection by 10% NaClO for 10 min. The same below

Fig. 3 Disinfection effect of PPM soaking time on onion seeds

Fig. 4 PCR amplification of fungal and bacterial strains

Table 1 Inhibition rates of different concentrations of PPM in four fungi

浓度 Concentration/（mL·L^-1）	S. vesicarium	A. alternata	F. proliferatum	F. oxysporum
0.25	65.71±5.86a	40.13±4.06a	49.05±3.33a	51.87±3.14a
0.50	72.14±6.84ab	53.62±2.63b	57.87±2.58ab	59.7±7.48ab
1.00	78.08±3.63abc	58.52±3.87b	60.36±2.49b	61.09±6.44ab
2.00	81.6±3.52bcd	62.39±3.07bc	63.25±5.18b	67.46±6.76ab
4.00	91.07±2.52cd	71.16±1.78c	73.74±3.17c	71.67±6.5bc
8.00	92.01±3.43cd	85.14±1.97d	79.96±1.31c	86.44±1.99c
16.00	94.97±2.6d	89.45±2.31d	81.65±2.13c	87.19±3.02c

Fig. 5 Inhibitory effects of different concentrations of PPM on Atlantibacter hermannii A：Bacteriostatic effect photograph. B：Diameter and significance of bacteriostatic circle, different lower-case letters indicate significant differences（P<0.05）and bacteriostatic circle with diameter ≤5 mm indicates no bacteriostatic effect

Fig. 6 Effects of PPM concentrations（mL/L）on the growth of onion cotyledons and roots

Fig. 7 Application of the optimal scheme in onion seed disinfection and sterile seedling culture A-B：Germination rate and contamination rate of 4 replicates. C-E：Aseptic seedling in 6, 10 and 20 d

Fig. 8 Application of optimal scheme in disinfection of three types of onion seeds and culture of aseptic seedling A-F：‘TIANZHENG201’（6 d）, ‘YY18’（6 d）, ‘GCH’（6 d）, ‘YOUYUE’（12 d）, ‘JIAMEI’（12 d）, ‘KAMUYI-153’（12 d）

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