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Acta Agronomica Sinica ›› 2022, Vol. 48 ›› Issue (1): 99-107.doi: 10.3724/SP.J.1006.2022.04275


NtPHYB1 interacts with light and temperature signal to regulate seed germination in Nicotiana tabacum L.

LI Zhen-Hua1,*(), WANG Xian-Ya1, LIU Yi-Ling2, ZHAO Jie-Hong3   

  1. 1College of Agriculture, Guizhou University, Guiyang 550025, Guizhou, China
    2Key Laboratory of Plant Resource Conservation and Germplasm Innovation in Mountainous Region (Ministry of Education), Guizhou University, Guiyang 550025, Guizhou, China
    3CNTC Key Laboratory of Molecular Genetics, Guizhou Academy of Tobacco Science, Guiyang 550081, Guizhou, China
  • Received:2020-12-15 Accepted:2021-04-14 Online:2022-01-12 Published:2021-06-28
  • Contact: LI Zhen-Hua E-mail:lixing_19841014@126.com
  • Supported by:
    National Natural Science Foundation of China(31860420);National Natural Science Foundation of China(32060512);Natural Science Foundation of Guizhou Province (QKHPT)([2019]1069);Science and Technology Plan Project of Guizhou Province (QKHJC)([2018]5781号);High-level Talents Introduction Project of Guizhou University (GDRJHZ)([2018]37号)


Temperature and light are two important environmental factors affecting seed germination, and phytochromes are the sensors of light and temperature. It is of great significance to investigate the interaction of phytochromes with light and temperature to regulate seed germination for guiding crop sowing. The aim of this study is to explore how the NtPHYB1 genotype regulates seed germination in response to change in light and temperature environments in tobacco. The average germination frequency of WT seeds was significantly higher than that of NtPHYB1-OE and NtPHYB1-RNAi seeds under 15 environments. The germination of NtPHYB1-OE seeds was inhibited by continuous light exposure, while the germination of NtPHYB1-RNAi seeds was repressed by darkness. At 15℃, the germinations of all three genotypic seeds were inhibited by the low-temperature, and the germination frequency of NtPHYB1-OE seeds was significantly lower than that of WT and NtPHYB1-RNAi seeds; while there was no significant effect in light signal. At 20℃ and 25℃, the temperature signal promoted the ability of seed germination, and the signal of light was dispensable, resulting in the highest germination frequencies for NtPHYB1-OE and WT seeds. At 30℃ and 35℃, the light signal was indispensable to maintain seed germination for all three genotypic seeds, resulting in the highest germination frequency for NtPHYB1-RNAi seeds. To sum up, there were interactions among NtPHYB1, photoperiod, and temperature in the regulation of seed germination in tobacco. The germination frequencies of tobacco seeds could be significantly improved by modifying NtPHYB1 gene or improving the cultivation environment.

Key words: seed germination, photothermal sensitivity, phytochrome, photothermal signal, natural selection, seasonal variation

Table 1

Effects of genotype, light, temperature and their interactions on seed germination in tobacco"

Genotype (G)

Light treatment (L)
(T, ℃)
Mean value (L×T×G)
Sample number
NtPHYB1过表达 持续黑暗Dark 15 17.78 qrstu NOP 8.36 9
NtPHYB1-OE 20 72.00 abcdefg ABCDE 9.26 9
25 78.00 abcdef ABC 7.91 9
30 26.78 opqr LMNO 13.14 9
35 2.89 u Q 2.62 9
NtPHYB1 RNA干扰 15 31.83 nopq KLMN 4.92 6
NtPHYB1-RNAi 20 65.67 cdefghi BCDEFG 7.50 6
25 75.00 abcdef ABCD 10.12 6
30 46.00 lmn GHIJKL 8.67 6
35 5.00 tu Q 1.41 6
野生型WT 15 38.00 mno IJKLM 6.93 3
20 85.67 ab AB 2.08 3
25 80.67 abc ABC 8.14 3
30 62.00 fghij CDEFGH 3.46 3
35 5.00 tu Q 2.00 3
NtPHYB1过表达 12 h光照/12 h 黑暗 15 16.22 rstu NOP 10.97 9
NtPHYB1-OE 12 h light/12 h dark 20 48.33 klm FGHIJK 15.36 9
Genotype (G)

Light treatment (L)
(T, ℃)
Mean value (L×T×G)
Sample number
25 63.33 efghi CDEFGH 10.55 9
30 74.11 abcdef ABCD 6.41 9
35 73.67 abcdef ABCD 10.32 9
NtPHYB1 RNA干扰 15 18.83 qrst MNOP 5.12 6
NtPHYB1-RNAi 20 55.00 hijkl DEFGHI 7.90 6
25 64.17 defghi CDEFGH 11.96 6
30 78.50 abcde ABC 9.67 6
35 79.33 abcde ABC 4.37 6
野生型WT 15 21.67 pqrs MNOP 4.93 3
20 69.33 cdefgh ABCDE 7.51 3
25 75.00 abcdef ABCD 2.65 3
30 79.67 abcd ABC 4.51 3
35 87.33 aA 3.06 3
NtPHYB1过表达 持续光照Light 15 8.67 stu OP 6.02 9
NtPHYB1- OE 20 38.11 mno IJKLM 18.31 9
25 53.11 jkl EFGHIJ 22.13 9
30 57.22 ghijk DEFGHI 10.93 9
35 65.00 cdefghi BCDEFG 17.68 9
NtPHYB1 RNA干扰 15 20.67 pqrs MNOP 8.98 6
NtPHYB1-RNAi 20 44.83 lmn HIJKL 7.88 6
25 68.33 cdefghi ABCDEF 8.07 6
30 70.33 bcdefgh ABCDE 11.33 6
35 66.17 cdefghi BCDEF 9.79 6
野生型WT 15 11.33 stu OP 3.51 3
20 34.00 mnop JKLMN 5.29 3
25 65.00 cdefghi BCDEFG 2.65 3
30 74.00 abcdef ABCDE 3.00 3
35 70.67 bcdefgh ABCDE 6.43 3

Table 2

Comparisons of mean difference within the genotype, light, and temperature treatment"

I J 平均差
Mean difference (I-J)
基因型Genotype NtPHYB1过表达NtPHYB1-OE NtPHYB1 RNA干扰NtPHYB1-RNAi -6.30***
野生型WT -10.94***
野生型WT -4.64*
野生型WT NtPHYB1过表达NtPHYB1-OE 10.94***
光Light 持续黑暗Dark 12 h光照/12 h黑暗 12 h light/12 h dark -14.70***
持续光照Light -5.04**
I J 平均差
Mean difference (I-J)
12 h光照/12 h黑暗 12 h light/12 h dark 持续黑暗Dark 14.70***
持续光照Light 9.66***
持续光照Light 持续黑暗Dark 5.04**
12 h光照/12 h黑暗 12 h light/12 h dark -9.66***
温度Temperature 15℃ 20℃ -36.31***
25℃ -48.74***
30℃ -41.00***
35℃ -30.39***
20℃ 15℃ 36.31***
25℃ -12.43***
30℃ -4.69*
35℃ 5.93**
25℃ 15℃ 48.74***
20℃ 12.43***
30℃ 7.74***
35℃ 18.35***
30℃ 15℃ 41.00***
20℃ 4.69*
25℃ -7.74***
35℃ 10.61***
35℃ 15℃ 30.39***
20℃ -5.93**
25℃ -18.35***
30℃ -10.61***

Table 3

Multi-factor variance analysis of the interactions among NtPHYB1, light, and temperature in the regulation of seed germination in tobacco"

III 型平方和
III sum of squares
基因型Genotype (G) 4800.759 2 2400.380 22.199*** 0.000
光处理Light (L) 7936.154 2 3968.077 36.697*** 0.000
温度处理Temperature (T) 63,436.466 4 15,859.116 146.666*** 0.000
G×L 975.230 4 243.807 2.255 0.064
G×T 1206.704 8 150.838 1.395 0.200
L×T 61,711.869 8 7713.984 71.339*** 0.000
G×L×T 3070.141 16 191.884 1.775* 0.036


Variation of seed germination under fixed temperature with photoperiodic change (A) and fixed photoperiodic with temperature change (B) in tobacco"

Table 4

NtPHYB1 regulates seed germination in response to signals of light and temperature"

Germination ratio among different genotype (G)
Environmental combinations of light and temperature
No significant difference
NtPHYB1-OE, WT and NtPHYB1-RNAi: T15L12, T15L24, T20L24, T25L0, T25L12, T30L12, T35L0, T35L12, and T35L24.
Significant difference
WT>NtPHYB1-OE: T15L0, T20L12, T25L24,T30L0, T30L24; WT>NtPHYB1-RNAi: T20L0 and T30L0; NtPHYB1-RNAi>NtPHYB1-OE: T25L24.
NtPHYB1-OE: T20L0, T25L0, T30L12, T35L12; WT: T20L0, T25L0, T25L12, T30L12, T30L24, T35L12; NtPHYB1-RNAi: T30L12, T30L24, T35L12.
NtPHYB1-OE: T15L0, T15L12, T15L24, T35L0; WT: T15L24, T35L0; NtPHYB1-RNAi: T35L0

Fig. 2

Diagram of phytochrome B integrates light and temperature signal to regulate seed germination in different seasons The pink block are seeds exposed to the ground, while the grey blocks are seeds buried underground. represents quick, slow, and obstructive conversions. In winter and early spring (<15℃), the low-temperature signal sensed by phytochrome B inhibited seeds germination, and the effect of the light signal was almost ineffective. During late spring and autumn (~20℃), temperature signal initiated germination with the combination of the light. In summer and early autumn (>30℃), light is indispensable to maintain seed germination by persistently activating the PHYB."

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