Silencing effect of different inoculation methods on differentially expressed genes in root and leaf of tobacco

doi:10.16178/j.issn.0528-9017.20240176

Abstract

Abstract:

The expression of different genes in different tissues of tobacco is different, and whether the inoculation method using TRV induced gene silencing (VIGS) technology in tobacco is suitable remains to be determined. Leaf injection method and root inoculation method were used to silence NtBBL and NtPPO7 differentially expressed genes in root and leaf of Yunyan 87, respectively, to determine the silencing efficiency of different inoculation methods and the effects of systematic analysis. The results showed that the relative expression level of NtBBL gene in the root was significantly higher than that in the leaf, and the relative expression level of NtPPO7 gene in the leaf was significantly higher than that in the root, which met the needs of the experiment. For silencing genes in root and leaf, leaf injection was suitable for silencing in leaf, and root inoculation was suitable for silencing in root. Leaf injection method has a higher silencing efficiency for high expression genes in leaves, while root inoculation method has a higher silencing efficiency for high expression genes in roots. BBL and PPO activity in tobacco were affected to a certain extent by different inoculation methods after silencing genes at high expression sites. The PPO activity in leaf was decreased by leaf injection method, while BBL activity in root was decreased by root inoculation method. Root inoculation method was more effective in reducing the nicotine content of tobacco leaves. It is necessary to select appropriate inoculation methods for the highly expressed genes in different tissues of tobacco, and different inoculation methods have different silencing effects on different genes. This study provides support for the application of VIGS technology in tobacco.

Key words: VIGS, tobacco, inoculation method, gene expression, enzyme activity

CLC Number:

S572

LI Wei, YANG Zaijun, XU Yanbiao, XIE Ke, XU Bingcong, WU Jianping, YE Xingrong, ZHOU Jianjun, ZU Qiongyao, ZHENG Cong. Silencing effect of different inoculation methods on differentially expressed genes in root and leaf of tobacco[J]. Journal of Zhejiang Agricultural Sciences, 2025, 66(5): 1151-1157.

Figures/Tables 8

Table 1 Test treatment

处理	侵染液
CK	不接种侵染液
CK1	叶片注射接种含pTRV2的侵染液(空载体对照)
CK2	灌根接种含pTRV2的侵染液(空载体对照)
A1	叶片注射接种含pTRV2-NtBBL侵染液
A2	灌根接种含pTRV2-NtBBL侵染液
B1	叶片注射接种含pTRV2-NtPPO7侵染液
B2	灌根接种含pTRV2-NtPPO7侵染液

Table 2 Primers for RT-qPCR

引物名称	引物序列(5'- 3')
NtBBL -F NtBBL -R	TTGCCAAGGTAAGTGACGTTC CCGTCCATCAGCATCAATAAG
NtPPO7 -F NtPPO7 -R	GCACGTTCAGTTCCTCCAGTTA TCGCTGATCGATGTTCAGTTC
26S rRNA-F 26S rRNA-R	GAAGAAGGTCCCAAGGGTTC TCTCCCTTTAACACCAACGG

Fig.1 Relative expression levels of NtBBL and NtPPO7 in roots and leaves

Fig.2 Relative gene expression of NtBBL in root and leaf treated by different inoculation methods

Fig.3 Relative gene expression of NtPPO7 in root and leaf treated by different inoculation methods

Fig.4 BBL activity in tobacco root treated with different inoculation methods

Fig.5 PPO activity in tobacco leaf treated with different inoculation methods

Fig.6 Nicotine content in tobacco leaf after NtBBL gene silencing by different inoculation methods

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