Prediction of VIGS efficiency by the Sfold program and its reliability analysis in Gossypium hirsutum
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- 作者：Xiaoyang Ge ; Jie Wu ; Chaojun Zhang ; Qianhua Wang ; Yuxia Hou…
- 关键词：VIGS ; Sfold program ; siRNA ; Target sequences ; Gossypium hirsutum
- 刊名：Chinese Science Bulletin
- 出版年：2016
- 出版时间：April 2016
- 年：2016
- 卷：61
- 期：7
- 页码：543-551
- 全文大小：995 KB
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Jie Wu (1)
Chaojun Zhang (1)
Qianhua Wang (1)
Yuxia Hou (3)
Zuoren Yang (1)
Zhaoen Yang (1)
Zhenzhen Xu (1)
Ye Wang (1)
Lili Lu (1)
Xueyan Zhang (1)
Jinping Hua (2)
Fuguang Li (1)
1. State Key Laboratory of Cotton Biology, Institute of Cotton Research, Chinese Academy of Agricultural Sciences, Anyang, 455000, China
2. Department of Plant Genetics and Breeding, Key Laboratory of Crop Heterosis and Utilization of Ministry of Education, Beijing Key Laboratory of Crop Genetic Improvement, China Agricultural University, Beijing, 100193, China
3. College of Science, China Agricultural University, Beijing, 100193, China - 刊物主题：Science, general; Life Sciences, general; Physics, general; Chemistry/Food Science, general; Earth Sciences, general; Engineering, general;
- 出版者：Springer Berlin Heidelberg
- ISSN：1861-9541
文摘
Genetic transformation in some plant species, including cotton (Gossypium hirsutum), is hampered by laborious and time-consuming processes and often unachievable. Virus-induced gene silencing (VIGS) by double-stranded RNAs can serve as a reverse-genetics tool to determine gene function. However, knockdown levels vary greatly when using a tobacco rattle virus-based vector that carries different cDNA fragments of a gene. How to choose the optional target fragment for high interference efficiency is very challenging. Addressing this challenge requires increasing the efficacy of small interference RNA (siRNA) in target fragment. Here, we describe a method to assess VIGS efficiency by comparing the following parameters of siRNA in target sequence: the disruption energy of the target (ΔGdisruption), the differential stability of siRNA duplex ends (DSSE), and the internal stability at positions 9–14 of the siRNA antisense strand (AIS), which are calculated by Sfold program (http://sfold.wadsworth.org). We find that the siRNAs with low ΔGdisruption, high DSSE and high AIS have high activity and easily result in high VIGS efficiency by experimentally testing the actual knockdown levels of the four target genes, GhPDS, GhCLA1, GhAOS1, and GhCXE1 via choosing different target sequences for each gene. Therefore, the Sfold program can be used to analyze target sequences when carrying out VIGS design to increase gene-silencing effects in plants.