Method for the identification of synthetic cell-or tissue-specific transcriptional regulatory regions
AssigneeBaylor College of Medicine
InventorsRobert J. Schwartz, Eric M. Eastman, Xuyang Li, Jeff Nordstrom
Life of this patent
Estimating the term…
Abstract
The invention concerns making and evaluating synthetic regulatory regions for controlling gene expression. The invention features a method for identifying transcription factor binding sites and a method for evaluating the regulatory functions of synthetic regulatory regions.
Claims42
A method of identifying binding sites for transcription factors, comprising the step of:
- identifying the oligonucleotides in oligonucleotide-protein complexes formed between one or more proteins of a cellular or nuclear extract and any of a plurality of double-stranded oligonucleotide fragments in a mixture of said fragments and said extract wherein said complexes are separated from free oligonucleotides in said mixture using size exclusion chromatography;
- and
- wherein the presence of a said oligonucleotide in a said complex is indicative that said oligonucleotide comprises a said binding site.
A method for evaluating a putative cell- or tissue-specific transcriptional regulatory region, comprising the step of:
- determining whether a cell comprising said putative transcriptional regulatory region in a transcriptional regulatory position to a selective gene is selected under selective conditions, wherein said selection of said cell will only occur if said selective gene is expressed at a sufficiently high level in said cell and wherein said selective gene will only be expressed at said sufficiently high level if said putative transcriptional regulatory region is active in said cell.
A method of identifying cell- or tissue-specific synthetic regulatory regions, comprising the steps of:
- identifying the oligonucleotides in oligonucleotide-protein complexes formed between one or more proteins of a cellular or nuclear extract and any of a plurality of double-stranded oligonucleotide fragments in a mixture of said fragments and said extract wherein said complexes are separated from free oligonucleotides in said mixture using size exclusion chromatography;
- and
- wherein the presence of a said oligonucleotide in a said complex is indicative that said oligonucleotide comprises a said binding site.
- evaluating a putative cell- or tissue-specific transcriptional regulatory region comprising said binding site under a selection condition by determining whether a cell comprising said putative transcriptional regulatory region in a transcriptional regulatory position to a selective gene is selected under selective conditions, wherein said selection of said cell will only occur if said selective gene is expressed at a sufficiently high level in said cell and wherein said selective gene will only be expressed at said sufficiently high level if said putative transcriptional regulatory region is active in said cell.
Dependencies are inferred from the claim text.