Two-color Fluorescent Reporter for Alternative Pre-mRNA Splicing
High-throughput Dynamic range of applications, both in vitro and in vivo Increased specificity to separate out alternative splicing events from other cellular processes The inclusion of a second reporter system significantly widens detection range Enhanced versatility due to alternative exon cassette and florescent protein swapping
Monitor alternative splicing events in individual living cells, heterogeneous cell populations and live animals.
Researchers at UCLA have developed a reporter assay for use in the detection of compounds that modulate alternative splicing. This novel and robust two-color fluorescent system provides a means to distinguish between general changes in splicing, such as efficiency, accuracy and transcription/translation abnormalities, and a particular alternative splicing event.
Patent Number: US8580511B2
Application Number: US2008305764A
Inventor: Stoilov, Peter G. | Black, Douglas L.
Priority Date: 19 Jun 2006
Priority Number: US8580511B2
Application Date: 26 May 2010
Publication Date: 12 Nov 2013
IPC Current: C07H002102 | C07H002104 | C12N001500 | C12Q000168
US Class: 43500613 | 4353201 | 5360231 | 5360235
Assignee Applicant: The Regents of the University of California
Title: Two-color fluorescent reporter for alternative pre-mRNA splicing
Usefulness: Two-color fluorescent reporter for alternative pre-mRNA splicing
Summary: The alternative pre-mRNA splicing reporter construct comprising a promoter operably linked to a nucleic acid sequence encoding first and second fluorescent proteins is useful for in vivo or in vitro monitoring of alternative pre-mRNA splicing events or for high-throughput screening of compounds that affect alternative pre-mRNA splicing for use in basic research, drug screening or drug design.
Novelty: New alternative pre-mRNA splicing reporter construct comprising a promoter operably linked to a nucleic acid sequence encoding first and second fluorescent proteins, useful for monitoring alternative pre-mRNA splicing events
Chemical/Material
Chemical/Material Application
State Of Development The invention has been tested as a high-throughput application (96- and 384-well plates) with a variety of fully functioning prototypes. Preliminary experiments indicate that the assay scales well to 1536 well format. Its use at UCLA has already revealed several compounds that modulate alternative splicing and can lead to development of therapy for Spinal Muscular Atrophy. Background Prior to translation, transcription generates a precursor molecule (pre-mRNA) that contains both introns (intervening sequences) and exons (protein coding regions). Alternative splicing pathways vary the production of a mature mRNA strand by modifying the introns removed and the exons joined. Depending on the splice sites, these mRNA variances give rise to proteome diversity by changing the encoded protein structure, which in turn can affect ligand binding, allosteric regulation, protein localization, etc. Although mutations in splice signals account for 15% of genetic diseases caused by point mutations indicating a pressing need for research into the mechanisms controlling alternative splicing, experimental efforts to discover compounds targeting splicing are hampered by a lack of reliable, reproducible, and high-throughput techniques. Related Materials Additional Technologies by these Inventors Tech ID/UC Case 20132/2006-704-0 Related Cases 2006-704-0
USA
