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A simple artificial microRNA vector based on ath-miR169d precursor from Arabidopsis

文献类型： 外文期刊

作者： Liu, Chong 1 ; Zhang, Lan 1 ; Sun, Jie 2 ; Luo, Yanzhong 1 ; Wang, Ming-Bo 3 ; Fan, Yun-Liu 1 ; Wang, Lei 1 ;

作者机构： 1.Chinese Acad Agr Sci, Biotechnol Res Inst, Natl Key Facil Crop Gene Resources & Genet Improv, Beijing 100081, Peoples R China

2.Shihezi Univ, Coll Agr, Key Lab Oasis Ecol Agr BINTUAN, Shihezi 832003, Peoples R China

3.CSIRO Plant Ind, Canberra, ACT 2601, Australia

关键词： Molecular Genetics: Biochemistry and Molecular Biophysics

期刊名称：MOLECULAR BIOLOGY REPORTS （ 影响因子：2.316； 五年影响因子：2.357 ）

ISSN：

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收录情况： SCI

摘要： Artificial microRNA (amiRNA) is becoming a powerful tool for silencing genes in plants, and several amiRNA vectors have recently been developed based on the natural precursor structures of ath-miR159a, ath-miR164b, ath-miR172a, ath-miR319a and osa-miR528. In this study we generated a simple amiRNA vector (pAmiR169d) based on the structure of Arabidopsis miR169d precursor (pre-miR169d). Two unique restriction sites were created inside the stem region of pre-miR169d, which allows for the artificial miRNA sequences to be cloned as either similar to 80 bp synthetic oligonucleotides or PCR products. A beta-glucuronidase:green florescent protein fusion gene (GUS-GFP) was efficiently silenced in transient assays using a pAmiR169d-derived construct targeting the GUS-GFP sequence. 5'-RACE showed that the target GUS-GFP transcript was cleaved precisely at the expected position across nucleotides 10 and 11 of the artificial miRNA. Thus, pAmiR169d allows for both easy construction of artificial miRNA constructs and efficient silencing of target genes in plants.

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