# US 5,352,595 A · Myod regulatory region > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 5,352,595 A - Document ID: US5352595A - Status: Granted - Kind: A - Application number: 07/753,520 - Filed: 1991-09-03 - Granted: 1994-10-04 - Earliest claimed: 1991-09-03 - Assignees: Fred Hutchinson Cancer Research Center - Inventors: Stephen J. Tapscott; Harold M. Weintraub; Theodore D. Palmer - Web page: https://patentgrep.com/patent/US5352595A ## CPC - C07K 14/475: Growth factors; Growth regulators - C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00) ## Abstract Isolated DNA or RNA molecules capable of hybridizing under stringent conditions to the myoD regulatory region, its proximal promoter and distal enhancer regulatory regions, and regulatory elements within the proximal and distal regions for binding basic helix-loop-helix proteins, MyoD, and proteins binding at SP1, AP1, CAAT, M-CAT, CArG, and MEF sites in DNA. DNA or RNA expression vectors for introducing a gene into a cell under the regulatory control of the myoD regulatory region. Transduced or transfected pre-muscle cells, myocytes, or myoblasts. Methods of inducing a muscle phenotype in a non-muscle cell, positively selecting for a cells expressing MyoD, and negatively selecting for cells expressing MyoD. ## Claims ### Claim 1 An isolated DNA or RNA molecule comprising a myoD proximal regulatory region capable of hybridizing under stringent conditions to the nucleotide sequence from positions 590 to 883 in SEQ ID NO: 1. ### Claim 2 (depends on 1) The isolated DNA or RNA molecule of claim 1, consisting essentially of both a first nucleotide sequence shown from positions 732 to 722 in SEQ ID NO: 1 and a second nucleotide sequence shown at positions 802 to 837 in SEQ ID NO: 1. ### Claim 3 An isolated DNA or RNA molecule comprising a myoD distal regulatory region capable of hybridizing under stringent conditions to the nucleotide sequence from positions 1 to 1200 in SEQ ID NO: 2. ### Claim 4 (depends on 3) The isolated DNA or RNA molecule of claim 3, consisting essentially of the nucleotide sequence shown from positions 421 to 1200 in SEQ ID NO: 2. ### Claim 5 A DNA or RNA expression vector for introducing a gent into a cell, comprising a myoD regulatory region operably linked to a gene whereby transcription of the gene is controlled by the myoD regulatory region when the vector is stably integrated into the genome of the cell, wherein the myoD regulatory region comprises a distal regulatory region and a proximal regulatory region, wherein the distal regulatory region comprises DNA or RNA capable of hybridizing under stringent conditions to the nucleotide sequence shown from positions 421 to 1200 in SEQ ID NO: 2, and wherein the proximal regulatory region comprises DNA or RNA capable of hybridizing under stringent conditions to both a first nucleotide sequence shown from positions 732 to 772 in SEQ ID NO: 1 and a second nucleotide sequence shown at positions 802 to 837 in SEQ ID NO: 1. ### Claim 6 (depends on 5) A cell transduced or transfected with the expression vector of claim 5. ### Claim 7 (depends on 6) The cell of claim 6, wherein the cell is selected from the group consisting of: pre-muscle cells, muscle cells, myocytes, myoblasts, and cells genetically engineered to express MyoD protein. ### Claim 8 In a method of inducing a muscle phenotype in a non-muscle cell, comprising the step of introducing a MyoD coding sequence into the non-muscle cell, the improvement comprising operably linking a myoD regulatory region to the MyoD coding region whereby transcription of the MyoD coding region is controlled by the myoD regulatory region when the vector is introduced into the cell, wherein the myoD regulatory region comprises a distal regulatory region and a proximal regulatory region, wherein the distal regulatory region comprises DNA or RNA capable of hybridizing under stringent conditions to the nucleotide sequence shown from positions 421 to 1200 in SEQ ID NO: 2, and wherein the proximal regulatory region comprises DNA or RNA capable of hybridizing under stringent conditions to both a first nucleotide sequence shown from positions 732 to 772 in SEQ ID NO: 1 and a second nucleotide sequence shown at positions 802 to 837 in SEQ ID NO: 1. ### Claim 9 A method of positively selecting for muscle cells, comprising: constructing a DNA or RNA expression vector comprising a myoD proximal regulatory region and a myoD distal regulatory region both operably linked to a marker gene whereby transcription of the marker gene is controlled by the myoD proximal and distal regulatory regions, introducing the expression vector into the genomes of a population of cells, and positively selecting for cells that express the marker, wherein the myoD proximal regulatory region is DNA or RNA capable of hybridizing under stringent conditions to the nucleotide sequence shown from positions 590 to 883 in SEQ ID NO: 1 and wherein the myoD distal regulatory region is DNA or RNA capable of hybridizing under stringent conditions to the nucleotide sequence shown from positions 1 to 1200 in SEQ ID NO: 2. ### Claim 10 A method of negatively selecting for a muscle cell, comprising: constructing a DNA or RNA expression vector comprising a myoD proximal regulatory region and a myoD distal regulatory region both operably linked to a gene encoding a toxin whereby transcription of the gene is controlled by the myoD proximal and distal regulatory regions, and introducing the vector into the genome of the cell, wherein the myoD proximal regulatory region is DNA or RNA capable of hybridizing under stringent conditions to the nucleotide sequence shown from positions 590 to 883 in SEQ ID NO: 1 and wherein the myoD distal regulatory region is DNA or RNA capable of hybridizing under stringent conditions to the nucleotide sequence shown from positions 1 to 1200 in SEQ ID NO: 2. ### Claim 11 A method of negatively selecting for muscle cells, comprising: introducing a DNA or RNA expression vector into the genomes of a population of cells, the expression vector comprising a myoD proximal regulatory region and a myoD distal regulatory region both operably linked to a gene encoding a protein for converting a prodrug to a toxic drug whereby transcription of the gene is controlled by the myoD proximal and distal regulatory regions, and contacting the cells with the prodrug, wherein the myoD proximal regulatory region is DNA or RNA capable of hybridizing under stringent conditions to the nucleotide sequence shown from positions 590 to 883 in SEQ ID NO: 1 and wherein the myoD distal regulatory region is DNA or RNA capable of hybridizing under stringent conditions to the nucleotide sequence shown from positions 1 to 1200 in SEQ ID NO: 2. ### Claim 12 An isolated DNA or RNA molecule comprising a myoD distal regulatory region capable of hybridizing under stringent conditions to the nucleotide sequence from positions 1 to 1978 in SEQ ID NO: 2. ### Claim 13 (depends on 12) The isolated DNA or RNA molecule of claim 12, consisting essentially of the nucleotide sequence shown from positions 421 to 1978 in SEQ ID NO: 2.