Rice repetitive DNA sequence RRD3: a plant promoter and its application to RNA interference.
Previously, a moderately repetitive DNA sequence (RRD3) was cloned from rice (Oryza sativa L.) by DNA renaturation kinetics. Sequence analysis revealed several conserved promoter motifs, including four TATA-boxes and a CAAT-box, and promoter activity was shown in Escherichia coli and mammalian expression systems. Here, we inserted the RRD3 fragment into the plant promoter-capture vector, pCAMBIA1391Z, and examined whether the RRD3 fragment has promoter activity in plants. Transgenic tobacco and rice calli both showed beta-glucuronidase (GUS) activity, indicating that RRD3 can act as a promoter in both monocot and dicot plants. Based on the promoter characteristic of RRD3, we designed a plant universal binary vector, pCRiRRD3, which is suitable for performing researches on plant RNA interference. This vector has two multiple cloning sites to facilitate sense and antisense cloning of the target sequence, separated by an intron fragment of 200 bp. The efficiency of the vector for gene silencing was assayed by histochemical and quantitative fluorometric GUS assays in transgenic tobacco. These research results suggested that this plant RNAi vector pCRiRRD3 can effectively perform gene silencing researches on both monocot and dicot plants.
Zhong
Jian
J
National Engineering Center of Cell Products, Amcellgene Co., Ltd., Tianjin 300457, China. zhongjianzhj@yahoo.com.cn
Wang
Hongbin
H
Zhang
Dangquan
D
Liu
Bing
B
Wang
Jinfa
J
D001483:Base Sequence; D017796:Caulimovirus / Q000235:genetics; D005822:Genetic Vectors; D012275:Oryza / Q000235:genetics*; D030821:Plants, Genetically Modified / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D034622:RNA Interference / Q000502:physiology*; D012091:Repetitive Sequences, Nucleic Acid / Q000502:physiology*; D014026:Nicotiana / Q000235:genetics*