A highly efficient method for construction of rice artificial MicroRNA vectors.

Abstract:

:Artificial microRNA (amiRNA) has become a powerful tool for gene silencing in plants. A new method for easy and rapid construction of rice artificial miRNA vector is described. The procedure involved modification of the pCAMBIA1300-UR vector by insertion of a 'vector modification fragment'. This was prepared from the precursor of Os-amiR528 by eliminating the central miRNA-containing region while simultaneously creating an AfeI restriction site. The fragment was then introduced to the destination vector to produce a multipurpose 'Highly Efficient gene Silencing Compatible vector' (HESC vector). AfeI was used to produce linearized HESC vectors, and a blunt end PCR product that included amiRNA sequence was cloned into this site by a single ligation reaction to create the completed amiRNA vector. Tests showed that the method was highly efficient, and greatly reduced the time needed for vector construction and resulted in a DNA sequence identical to that of the current method, making it particularly suitable for use in a systems biology approach to functional genomic research.

journal_name

Mol Biotechnol

journal_title

Molecular biotechnology

authors

Wang X,Yang Y,Yu C,Zhou J,Cheng Y,Yan C,Chen J

doi

10.1007/s12033-010-9291-4

subject

Has Abstract

pub_date

2010-11-01 00:00:00

pages

211-8

issue

3

eissn

1073-6085

issn

1559-0305

journal_volume

46

pub_type

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