A novel approach for the construction of plant amiRNA expression vectors

Yan, Hong; Deng, Xuan; Cao, Yanwei; Huang, Junyan; Ma, Lixin; Zhao, Bin

doi:10.1016/j.jbiotec.2010.10.078

Cited by 15 publications

(3 citation statements)

References 24 publications

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“…However, the production of the miR319-based amiRNA required multiple PCR reactions on the miR319 backbone. Previously, we and others evaluated the effectiveness of a truncated, 120-nt long, osa-MIR528 sequence for the production of an amiRNA [11,38]. Hence, in the present study, the miR319 backbone was replaced with the smaller, osa-MIR528 backbone, which requires only a single PCR for the construction of the amiRNA.…”

Section: Discussionmentioning

confidence: 99%

Alterations of the viroid regions that interact with the host defense genes attenuate viroid infection in host plant

Adkar-Purushothama

Perreault

2018

RNA Biology

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Understanding in intimate details how the viroid interaction with host's defense genes is a cornerstone for developing viroid resistant plants. In this present study, small RNAs (sRNA) derived from Potato spindle tuber viroid (PSTVd) were studied in silico in order to detect any interactions with the serine threonine kinase receptor, a transmembrane protein that plays a role in disease resistance in plants. Using molecular biology techniques, it was determined that PSTVd infection negatively affects at least three serine threonine kinase receptors as well as with three other genes that are known to be involved in the overall development of the tomato plants. The transient expression of these putative PSTVd-sRNAs, using the microRNA sequence as a backbone, in tomato plants induced phenotypes similar to viroid infection. Mutants created by altering the sequence of PSTVd in these regions failed to infect the tomato plant. The data presented here illustrates the importance of these regions in viroid survival, and suggests a possible avenue of exploration for the development of viroid resistant plants.

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Section: Discussionmentioning

confidence: 99%

Alterations of the viroid regions that interact with the host defense genes attenuate viroid infection in host plant

Adkar-Purushothama

Perreault

2018

RNA Biology

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“…The central region (14B-15B), containing a fragment of the gene target sequence, was constructed using gene-specific oligonucleotides, as described in Supplemental Figure S7B. In order to validate the proposed structure, Arabidopsis PDS silencing was assayed using a gene target fragment that was formerly described by Yan et al (2011;Supplemental Fig. S7E).…”

Section: Silencing Toolsmentioning

confidence: 99%

GoldenBraid 2.0: A Comprehensive DNA Assembly Framework for Plant Synthetic Biology

et al. 2013

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Plant synthetic biology aims to apply engineering principles to plant genetic design. One strategic requirement of plant synthetic biology is the adoption of common standardized technologies that facilitate the construction of increasingly complex multigene structures at the DNA level while enabling the exchange of genetic building blocks among plant bioengineers. Here, we describe GoldenBraid 2.0 (GB2.0), a comprehensive technological framework that aims to foster the exchange of standard DNA parts for plant synthetic biology. GB2.0 relies on the use of type IIS restriction enzymes for DNA assembly and proposes a modular cloning schema with positional notation that resembles the grammar of natural languages. Apart from providing an optimized cloning strategy that generates fully exchangeable genetic elements for multigene engineering, the GB2.0 toolkit offers an evergrowing open collection of DNA parts, including a group of functionally tested, premade genetic modules to build frequently used modules like constitutive and inducible expression cassettes, endogenous gene silencing and protein-protein interaction tools, etc. Use of the GB2.0 framework is facilitated by a number of Web resources that include a publicly available database, tutorials, and a software package that provides in silico simulations and laboratory protocols for GB2.0 part domestication and multigene engineering. In short, GB2.0 provides a framework to exchange both information and physical DNA elements among bioengineers to help implement plant synthetic biology projects.

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“…Several studies have reported a simplified method for amiRNA vector construction but have either modified the amiRNA skeleton or used a different amiRNA skeleton (see, for example, (Liu et al 2010)). Some other studies describe a different strategy to generate amiRNAs but require more experimental steps than desirable (Yan et al 2011a;Yan et al 2011b). Our method (summarized in Fig.…”

Section: Introductionmentioning

confidence: 99%

Two-step method for constructing Arabidopsis artificial microRNA vectors

et al. 2012

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Artificial microRNA (amiRNA) technology is used for gene silencing in Arabidopsis. We describe a method for constructing amiRNA vectors that requires only one PCR and one ligation reaction. Vectors produced by this method are the same as those from the method of Schwab et al. (Plant Cell 2006, 18:1121-1133). Transgenic plants created by this method can therefore be tested in the same way or compared with existing transgenic material without the risk of alteration to the amiRNA skeleton. With optimized parameters, 36-42 % colonies had the insertion in the expected orientation and 85-95 % of these had the correct sequence. Using this method, a transient gene knock-down analysis in Arabidopsis could be completed in 4-5 days.

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A novel approach for the construction of plant amiRNA expression vectors

Cited by 15 publications

References 24 publications

Alterations of the viroid regions that interact with the host defense genes attenuate viroid infection in host plant

Alterations of the viroid regions that interact with the host defense genes attenuate viroid infection in host plant

GoldenBraid 2.0: A Comprehensive DNA Assembly Framework for Plant Synthetic Biology

Two-step method for constructing Arabidopsis artificial microRNA vectors

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