Synthesis of tagged nucleic acids

US8329887B2 · kind B2 · utility

Assignee

• CELLSCRIPT, LLC · US

Inventors

• Gary Dahl · Madison, US
• Roy Rabindranauth Sooknanan · Beaconsfield, CA

Key dates

• Filing date: Sep 20, 2011
• Grant date: Dec 11, 2012
• Expiry date: Sep 20, 2031

Classification

• Technology area (CPC C): Chemistry; Metallurgy
• CPC primary: C12Q1/6865
• WIPO field: Biotechnology
• WIPO sector: Chemistry

Abstract

The present invention relates generally to methods, compositions and kits for synthesizing sense RNA molecules from one or more RNA molecules of interest in a sample. In exemplary embodiments, the methods use a terminal tagging oligoribonucleotide (rTTO) to join a DNA sequence tag to the 3′-termini of first-strand cDNA molecules. The use of an rTTO comprising ribonucleotides results in decreased oligonucleotide-derived background synthesis of RNA in the absence of sample RNA and, surprisingly and unexpectedly, also results in significantly increased yields of sense RNA molecules that exhibit sequences that are substantially identical to those of the RNA molecules of interest in the sample. The sense RNA molecules also have an RNA sequence tag on their 5′-termini that is useful for fixing the lengths of sense RNA molecules that are synthesized in a second or subsequent round.