Evaluation and identification of candidate genes for artificial microRNA-mediated resistance to tomato spotted wilt virus

Mitter, Neena, Zhai, Ying, Bai, Anh Xu, Chua, Keith, Eid, Sahar, Constantin, Myrna, Mitchell, Roger and Pappu, Hanu R. (2016) Evaluation and identification of candidate genes for artificial microRNA-mediated resistance to tomato spotted wilt virus. Virus Research, 211 151-158. doi:10.1016/j.virusres.2015.10.003


Author Mitter, Neena
Zhai, Ying
Bai, Anh Xu
Chua, Keith
Eid, Sahar
Constantin, Myrna
Mitchell, Roger
Pappu, Hanu R.
Title Evaluation and identification of candidate genes for artificial microRNA-mediated resistance to tomato spotted wilt virus
Journal name Virus Research   Check publisher's open access policy
ISSN 0168-1702
1872-7492
Publication date 2016-01-04
Year available 2015
Sub-type Article (original research)
DOI 10.1016/j.virusres.2015.10.003
Open Access Status Not Open Access
Volume 211
Start page 151
End page 158
Total pages 8
Place of publication Amsterdam, Netherlands
Publisher Elsevier
Collection year 2016
Language eng
Formatted abstract
Tomato spotted wilt virus (TSWV) is an economically important viral pathogen of a wide range of field and horticultural crops. We developed an artificial microRNA (amiRNA) strategy against TSWV, targeting the nucleoprotein (N) and silencing suppressor (NSs) genes. The amiRNA constructs replaced the natural miRNA in a shortened Arabidopsis 173-nucleotide (nt) miR159a precursor backbone (athmiR159a) withoutthe stem base extending beyond the miR/miR* duplex. Further, each amiRNA was modified to contain a mismatch (wobble) sequence at nucleotide position 12 and 13 on the complementary strand amiRNA*, mimicking the endogenous miR159a sequence structure. Transient expression in Nicotiana benthamiana demonstrated that the introduction of a wobble sequence did not alter amiRNA expression levels. Following challenge inoculation with TSWV, plants expressing N-specific amiRNAs with or without the wobble remained asymptomatic and were negative for TSWV by ELISA. In contrast, plants expressing the NSs-specific amiRNAs were symptomatic and accumulated high levels of TSWV. Similar findings were obtained in stably transformed Nicotiana tabacum plants. Our results show that a shortened 173-nt athmiR159a backbone is sufficient to express amiRNAs and that the presence of mismatch at position 12–13 does not influence amiRNA expression or conferring of resistance. We also show that selection of target gene and positional effect are critical in amiRNA based approach for introducing resistance. These findings open the possibility of employing the amiRNA approach for broad spectrum resistance to tospoviruses as well as other viruses.
Keyword RNA silencing
Tomato spotted wilt virus
Artificial microRNA
Virus resistance
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Queensland Alliance for Agriculture and Food Innovation
Official 2016 Collection
 
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Created: Sun, 13 Dec 2015, 22:33:06 EST by Dr Neena Mitter on behalf of Centre for Plant Science