Two-step method for constructing Arabidopsis artificial microRNA vectors

Wang, Xüming; Yang, Yong; Zhou, Jie; Chen, Yu; Cheng, Ye; Yan, Chengqi; Chen, Jianping

doi:10.1007/s10529-012-0901-z

Cited by 4 publications

(1 citation statement)

References 13 publications

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“…Nonlinearized plasmid molecules with no amiRNA insert fail to propagate in E. coli ccdB-sensitive strains, such as DH5a or DH10B. In summary, compared with other amiRNA cloning methods (Schwab et al, 2006;Qu et al, 2007;Chen et al, 2009;Molnar et al, 2009;Wang et al, 2010Wang et al, , 2012Eamens et al, 2011;Yan et al, 2011;Liang et al, 2012;Zhou et al, 2013), this method is relatively simple, fast, and cost effective (Fig. 2C).…”

Section: Atmir390a-based Vectorsmentioning

confidence: 92%

New Generation of Artificial MicroRNA and Synthetic Trans-Acting Small Interfering RNA Vectors for Efficient Gene Silencing in Arabidopsis

et al. 2014

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Section: Atmir390a-based Vectorsmentioning

confidence: 92%

New Generation of Artificial MicroRNA and Synthetic Trans-Acting Small Interfering RNA Vectors for Efficient Gene Silencing in Arabidopsis

et al. 2014

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Universal vectors for constructing artificial microRNAs in plants

Zhou

Chen

et al. 2013

Biotechnol Lett

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Universal amiRNA vectors (pUAs) for constructing plant amiRNAs in Arabidopsis and rice have been developed. By using type IIg restriction enzyme, BaeI, a single amiRNA construct can be produced using only one PCR and one ligation reaction. Thus, only one pair of primers is required for each amiRNA vector and these can be designed to be compatible with existing or newly developed methods. Because the BaeI recognition sequence is completely digested, there is no modification to the miRNA backbone, therefore avoids the risk of sequence changes that may affect downstream analysis. Based on these vectors, specific amiRNA constructs were created and verified. With optimized parameters, 38-45% colonies for each amiRNA construct contain insertions with the expected orientation, and approximately 80% of these colonies have the correct sequences.

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Artificial microRNA mediated gene silencing in plants: progress and perspectives

2014

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Homology based gene silencing has emerged as a convenient approach for repressing expression of genes in order to study their functions. For this purpose, several antisense or small interfering RNA based gene silencing techniques have been frequently employed in plant research. Artificial microRNAs (amiRNAs) mediated gene silencing represents one of such techniques which can utilize as a potential tool in functional genomics. Similar to microRNAs, amiRNAs are single-stranded, approximately 21 nt long, and designed by replacing the mature miRNA sequences of duplex within pre-miRNAs. These amiRNAs are processed via small RNA biogenesis and silencing machinery and deregulate target expression. Holding to various refinements, amiRNA technology offers several advantages over other gene silencing methods. This is a powerful and robust tool, and could be applied to unravel new insight of metabolic pathways and gene functions across the various disciplines as well as in translating observations for improving favourable traits in plants. This review highlights general background of small RNAs, improvements made in RNAi based gene silencing, implications of amiRNA in gene silencing, and describes future themes for improving value of this technology in plant science.

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Two-step method for constructing Arabidopsis artificial microRNA vectors

Cited by 4 publications

References 13 publications

New Generation of Artificial MicroRNA and Synthetic Trans-Acting Small Interfering RNA Vectors for Efficient Gene Silencing in Arabidopsis

New Generation of Artificial MicroRNA and Synthetic Trans-Acting Small Interfering RNA Vectors for Efficient Gene Silencing in Arabidopsis

Universal vectors for constructing artificial microRNAs in plants

Artificial microRNA mediated gene silencing in plants: progress and perspectives

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