白背飞虱海藻糖合成酶基因调控几丁质合成的功能

doi:10.3864/j.issn.0578-1752.2019.19.007

Abstract

Abstract:

【Background】 It is well known that trehalose-6-phosphate synthase (TPS) plays an important role in trehalose synthesis, which can mediate trehalose metabolism to regulate chitin synthesis and insect development. 【Objective】 In this study, the effect of silencing SfTPS was detected, the molting status of Sogatella furcifera was observed and the content of chitin and the quantitative expression of chitin synthase (CHS) gene were determined through inhibiting the expression of TPS by RNAi in S. furcifera. The purpose is to explore the potential regulatory effects of SfTPS on the chitin synthesis in S. furcifera.【Method】 The S. furcifera population, which has been fed in laboratory for many years, was used as experimental material. The double-stranded RNA (dsRNA) of SfTPS and GFP were synthesized in vitro in order to study the potential function of TPSs, the RNA of two dsSfTPSs (dsSfTPS1 and dsSfTPS2) was injected into S. furcifera, respectively. Firstly, after dsRNA injection, the total RNA of S. furcifera was extracted by Trizol method at 48 h, and the first strand DNA was synthesized as the template of quantitative real-time PCR (qRT-PCR). As well as qRT-PCR was used to detect the silencing of SfTPS expression for the effect of RNAi. Secondly, the whole chitin of S. furcifera was detected at 48 and 72 h after dsRNA injection, and photographs were taken of the winged developmental malformations. Finally, the relative expression level of SfCHS in S. furcifera was detected by qRT-PCR, and the roles of SfTPS1 and SfTPS2 in the regulation of chitin synthesis were analyzed.【Result】 Compared with dsGFP injection, the expression of SfCHS increased after dsSfTPS1 and dsSfTPS2 injection, as well as the chitin content increased, which lead to the wing deformity of S. furcifera. The qRT-PCR results showed that the expression of SfTPS was significantly inhibited after dsRNA injection of a single SfTPS, which was less than 30% of that in the control group injected with dsGFP. In addition, the expression of another SfTPS also decreased significantly after a single SfTPS dsRNA injection. The adult wings of S. furcifera were all long wings after dsSfTPS1 and dsSfTPS2 injection. Some deformities such as wing curl in a certain ratio had been found in these long wings, and a certain ratio of mortality was found at 48 and 72 h. The chitin content increased significantly at 72 h after these two dsSfTPSs RNA injection. Compared with the control group, the expression of SfCHS1 and SfCHS1a increased significantly at 72 h after dsSfTPS1 injection, followed it increased significantly at 48 and 72 h after dsSfTPS2 injection. In the same time, the expression of SfCHS1b increased significantly after dsSfTPS1 and dsSfTPS2 injection.【Conclusion】 The SfTPS can control the synthesis of chitin through the regulation of chitin synthase gene in S. furcifera. The results are helpful to evaluate the regulatory role of SfTPS in S. furcifera and other insects and as the potential pest control target. It provides a theoretical basis for further development and screening of effective trehalose-6-phosphate synthase inhibitors to control S. furcifera pests and so on.

Key words: white-backed planthopper (Sogatella furcifera), trehalose-6-phosphate synthase (TPS), RNA interference, chitin synthesis, quantitative real-time PCR (qRT-PCR)

ZHANG DaoWei,YU YaYa,PAN BiYing,KANG Kui,ZENG BoPing,CHEN Jing,TANG Bin. Regulation Function of Trehalose-6-phosphate Synthase Genes on Chitin Synthesis in Sogatella furcifera[J].Scientia Agricultura Sinica, 2019, 52(19): 3357-3366.

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References 39

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引物名称 Primer name	上游引物 Forward primer (5′-3′)	下游引物 Reverse primer (5′-3′)	产物长度 Length (bp)
dsSfTPS1	CCCGTTGTGGTGAGAAATA	CAAGGTGGGAATGGAATG	473
dsSfTPS1-T7	T7-CCCGTTGTGGTGAGAAATA	T7-CAAGGTGGGAATGGAATG	523
dsSfTPS2	CGCATAGACCGCAACAAC	TCGCAACGGAGTAACCAG	459
dsSfTPS2-T7	T7-CGCATAGACCGCAACAAC	T7-TCGCAACGGAGTAACCAG	509
dsGFP	AAGGGCGAGGAGCTGTTCACCG	CAGCAGGACCATGTGATCGCGC	688
dsGFP-T7	T7-AAGGGCGAGGAGCTGTTCACCG	T7-CAGCAGGACCATGTGATCGCGC	738

引物名称 Primer name	上游引物 Forward primer (5′-3′)	下游引物 Reverse primer (5′-3′)	产物长度 Length (bp)
Q-SfTPS1	CCGATTCGCTACATCTACG	GACAAACTCTTTCGCCACTAA	123
Q-SfTPS2	GATGCTGAGGGCAAAGAC	TGTGGAAGCCGACAAAGT	226
Q-SfCHS1	GATTGGTCATTGGCTTCAGA	GTAATGTCTTGCTTCGTCAG	151
Q-SfCHS1a	CTTCGGTGTTTGGTTTCTT	TGGGTAACATCATCATAGGA	136
Q-SfCHS1b	GAGAAGGCGAGAATAGCA	GCAGCAAGAACACGATTA	103
Q-18S rRNA	GCCCCGTAATCGGAATGAGT	GACAAGACGTCCCGCAAAAC	205

Regulation Function of Trehalose-6-phosphate Synthase Genes on Chitin Synthesis in Sogatella furcifera

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