Single tube, high throughput cloning of inverted repeat constructs for double-stranded RNA expression

5Citations
Citations of this article
20Readers
Mendeley users who have this article in their library.

Abstract

BACKGROUND: RNA interference (RNAi) has emerged as a powerful tool for the targeted knockout of genes for functional genomics, system biology studies and drug discovery applications. To meet the requirements for high throughput screening in plants we have developed a new method for the rapid assembly of inverted repeat-containing constructs for the in vivo production of dsRNAs.<br /><br />METHODOLOGY/PRINCIPAL FINDINGS: The procedure that we describe is based on tagging the sense and antisense fragments with unique single-stranded (ss) tails which are then assembled in a single tube Ligase Independent Cloning (LIC) reaction. Since the assembly reaction is based on the annealing of unique complementary single stranded tails which can only assemble in one orientation, greater than ninety percent of the resultant clones contain the desired insert.<br /><br />CONCLUSION/SIGNIFICANCE: Our single-tube reaction provides a highly efficient method for the assembly of inverted repeat constructs for gene suppression applications. The single tube assembly is directional, highly efficient and readily adapted for high throughput applications.

Cite

CITATION STYLE

APA

Hauge, B., Oggero, C., Nguyen, N., Fu, C., & Dong, F. (2009). Single tube, high throughput cloning of inverted repeat constructs for double-stranded RNA expression. PLoS ONE, 4(9). https://doi.org/10.1371/journal.pone.0007205

Register to see more suggestions

Mendeley helps you to discover research relevant for your work.

Already have an account?

Save time finding and organizing research with Mendeley

Sign up for free