Establishment and Optimization of Virus-induced Gene Silencing System in Bougainvillea peruviana ‘Thimma’

doi:10.13560/j.cnki.biotech.bull.1985.2022-1266

Abstract

Abstract:

Bougainvillea peruviana ‘Thimma’, belonging to the family Nyctaginaceae, has high ornamental values in landscape. It is an ideal material to study the regulation of flower and leaf coloration. It is vital to establish a rapid and efficient gene verification system in B. peruviana. In this study, phytoene desaturase gene（PDS）was used as an indicator gene, we explored the effects of different inoculation sites and fragment length on the silencing effects of endogenous PDS mRNA in B. peruviana leaves induced by tobacco rattle virus（TRV）and established an optimized virus-induced gene silencing system（VIGS）for B. peruviana. The results showed that the silencing effect of leaf friction injection was more significant than that of tender stems friction injection. Additionally, PDS gene silencing was induced by both the inserted fragment of 336 bp and 457 bp, and the latter one was more significant. The VIGS system of B. peruviana is preliminarily constructed, which can be applied to the following gene function research.

Key words: Bougainvillea peruviana ‘Thimma’, phytoene desaturase（PDS）, tobacco rattle virus（TRV）, virus-induced gene silencing（VIGS）

LIU Zhen-yin, DUAN Zhi-zhen, PENG Ting, WANG Tong-xin, WANG Jian. Establishment and Optimization of Virus-induced Gene Silencing System in Bougainvillea peruviana ‘Thimma’[J]. Biotechnology Bulletin, 2023, 39(7): 123-130.

Figures/Tables 8

References 31

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引物 Primer	引物序列 Primer sequence（5'-3'）	长度 Length/bp
BpPDS336-EcoRI-F	TAAGGTTACCGAATTCTGCATTTTGATTGCTTTG	368
BpPDS336-EcoRI-R	GCCTTCTAGAGAATTCAATGACTGGAACCCCTACTA	368
BpPDS457-EcoRI-F	TAAGGTTACCGAATTCGGTTTGAGTGTCCAAGAGTG	489
BpPDS457-EcoRI-R	GCCTTCTAGAGAATTCTAATGACTGGAACCCCTACT	489
pTRV1-F	CCTGGTGCTCAGTACATAACTTTCC	257
pTRV1-R	TCTTTGCTTACATCGTCCTCTTTCA	257
pTRV2-F	ACGCTGTTTGAGGGAAAAGTAG	321
pTRV2-R	CCGATCAATCAAGATCAGTCGAGA	321
BpActin-RT-F	TATGAACTTCCTGATGGG	125
BpActin-RT-R	GAATTGTAGGTTGTCTCG	125
BpPDS-RT-F	ATTCGCCTCGTCCTGATA	103
BpPDS-RT-R	ATGGGTTTGTGACCTGCA	103

引物 Primer	引物序列 Primer sequence（5'-3'）	长度 Length/bp
BpPDS336-EcoRI-F	TAAGGTTACCGAATTCTGCATTTTGATTGCTTTG	368
BpPDS336-EcoRI-R	GCCTTCTAGAGAATTCAATGACTGGAACCCCTACTA	368
BpPDS457-EcoRI-F	TAAGGTTACCGAATTCGGTTTGAGTGTCCAAGAGTG	489
BpPDS457-EcoRI-R	GCCTTCTAGAGAATTCTAATGACTGGAACCCCTACT	489
pTRV1-F	CCTGGTGCTCAGTACATAACTTTCC	257
pTRV1-R	TCTTTGCTTACATCGTCCTCTTTCA	257
pTRV2-F	ACGCTGTTTGAGGGAAAAGTAG	321
pTRV2-R	CCGATCAATCAAGATCAGTCGAGA	321
BpActin-RT-F	TATGAACTTCCTGATGGG	125
BpActin-RT-R	GAATTGTAGGTTGTCTCG	125
BpPDS-RT-F	ATTCGCCTCGTCCTGATA	103
BpPDS-RT-R	ATGGGTTTGTGACCTGCA	103

处理 Treatment	叶绿素a含量 Chlorophyll a content/（mg·g^-1）	叶绿素b含量 Chlorophyll b content/（mg·g^-1）	叶绿素含量 Chlorophyll content/（mg·g^-1）	类胡萝卜素含量 Carotenoid content/（mg·g^-1）
空白对照	0.327 ± 0.021 a	0.214 ± 0.030 a	0.541 ± 0.052 a	0.114 ± 0.004 a
阴性对照	0.300 ± 0.007 a	0.153 ± 0.010 b	0.470 ± 0.025 a	0.114 ± 0.010 a
pTRV2-BpPDS336	0.230 ± 0.023 b	0.090 ± 0.010 c	0.310 ± 0.060 b	0.087 ± 0.003 b
pTRV2-BpPDS457	0.190 ± 0.016 c	0.053 ± 0.010 d	0.260 ± 0.035 b	0.086 ± 0.020 b

处理 Treatment	叶绿素a含量 Chlorophyll a content/（mg·g^-1）	叶绿素b含量 Chlorophyll b content/（mg·g^-1）	叶绿素含量 Chlorophyll content/（mg·g^-1）	类胡萝卜素含量 Carotenoid content/（mg·g^-1）
空白对照	0.327 ± 0.021 a	0.214 ± 0.030 a	0.541 ± 0.052 a	0.114 ± 0.004 a
阴性对照	0.300 ± 0.007 a	0.153 ± 0.010 b	0.470 ± 0.025 a	0.114 ± 0.010 a
pTRV2-BpPDS336	0.230 ± 0.023 b	0.090 ± 0.010 c	0.310 ± 0.060 b	0.087 ± 0.003 b
pTRV2-BpPDS457	0.190 ± 0.016 c	0.053 ± 0.010 d	0.260 ± 0.035 b	0.086 ± 0.020 b