Heterologous expression of artificial miRNAs from rice dwarf virus in transgenic rice

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• 作者：Runhong Sun (1) (2) (3)
  Peng Du (2)
  Lin Jiang (2)
  Derong An (1)
  Yi Li (2)
  
• 关键词：Expression efficiency ; Artificial miRNAs (amiRNAs) ; Copy number ; osa ; miRNA528 ; Rice dwarf virus (RDV)
• 刊名：Plant Cell, Tissue and Organ Culture
• 出版年：2014
• 出版时间：March 2014
• 年：2014
• 卷：116
• 期：3
• 页码：353-360
• 全文大小：2,064 KB
• 作者单位：Runhong Sun (1) (2) (3)
  Peng Du (2)
  Lin Jiang (2)
  Derong An (1)
  Yi Li (2)
  
  1. State Key Laboratory of Crop Stress Biology for Arid Areas and College of Plant Protection, Northwest A&F University, Yangling, Shaanxi, China
  2. Peking-Yale Joint Center for Plant Molecular Genetics and Agrobiotechnology, The State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing, China
  3. Henan Key Laboratory of Crop Pest Control of Henan Province, Key Laboratory of Pest Management in South of North China for Ministry of Agriculture of PRC, Institute of Plant Protection, Henan Academy of Agricultural Sciences, Zhengzhou, China
  
• ISSN：1573-5044

文摘

In contrast to hairpin RNAs, in which heterogeneous small RNAs are processed from double-stranded RNA to have potential off-target effects on endogenous other genes, artificial miRNAs (amiRNAs) have advantages of exquisite specificity and non-transitivity to thus target individual genes and groups of endogenous genes. Earlier studies showed that amiRNA engineering based on osa-miRNA528 precursor could efficiently trigger endogenous gene silencing and modulate agronomic traits in rice. However, both the expression efficiency of heterologous amiRNAs based on osa-miRNA528 precursor and the correlation of copy number with the relative expression level of amiRNAs remain unknown. In the present study, five amiRNAs (S9-1174, S9-1192, S11-864, S11-868 and S11-869) targeting different sites of S9 and S11 negative strands in rice dwarf virus (RDV) genome were constructed using endogenous osa-miRNA528 precursor as backbone. After identification by Northern blot, two amiRNAs (S9-1174 and S9-1192) targeting S9 negative strand in RDV genome were highly expressed, whereas in three tested amiRNAs targeting S11 negative strand in RDV genome, only two amiRNAs (S11-868 and S11-869) were processed efficiently. T0 generation transgenic rice containing amiRNAs (S9-1174, S9-1192, S11-868 and S11-869) exhibited different expression ratios of amiRNAs, accounting for 90.0, 90.0, 66.7 and 77.8?%, respectively. In addition, combination analysis with the relative amiRNA expression levels and its copy number revealed that the relative expression levels of amiRNAs had no relation to the copy number of T-DNA insert in transgenic rice.