Targeted RNA degradation using nuclear antisense RNA

An isolated mammalian cell containing a vector having incorporated therein a DNA construct comprising:

• a. a transcriptional promoter which provides efficient expression of an operably linked DNA segment such that inhibition of expression of a gene can be achieved with a 5-fold excess of antisense over sense RNA;
• b. a DNA segment operably linked to said transcriptional promoter;
• and
• c. sequences encoding a cis-ribozyme operably linked to said DNA segment,
• such that expression of the DNA segment produces a ribonucleotide sequence which is complementary to a ribonucleotide sequence transcribed from a gene, and said expressed ribonucleotide sequence is accumulated in the cell nucleus and inhibits the expression of said gene.

A method of inhibiting the expression of a gene in a mammalian cell comprising introducing into a mammalian cell a DNA construct comprising:

• a. a transcriptional promoter which provides efficient expression of an operably linked DNA segment such that inhibition of expression of a gene can be achieved with a 5-fold excess of antisense over sense RNA;
• b. a DNA segment operably linked to said transcriptional promoter;
• and
• c. sequences encoding a cis-ribozyme operably linked to said DNA segment,
• such that expression of the DNA segment produces a ribonucleotide sequence which is complementary to a ribonucleotide sequence transcribed from a gene, and said expressed ribonucleotide sequence is accumulated in the cell nucleus and inhibits the expression of said gene.

A method of inhibiting the expression of a gene in a mammalian cell, comprising the steps of:

• a. introducing into a mammalian cell a DNA construct comprising:
• i. a transcriptional promoter which provides efficient expression of an operably linked DNA segment such that inhibition of expression of a gene can be achieved with a 5-fold excess of antisense over sense RNA;
• ii. a DNA segment operably linked to said transcriptional promoter;
• and
• iii. sequences encoding a cis-ribozyme operably linked to said DNA segment,
• such that expression of the DNA segment produces a ribonucleotide sequence which is complementary to a ribonucleotide sequence transcribed from a gene, and said expressed ribonucleotide sequence is accumulated in the cell nucleus and inhibits the expression of said gene, whereby a transformed cell is obtained;
• and
• b. expressing the DNA segment to thereby inhibit the expression of said gene.

A DNA construct comprising:

• a. a human cytomegalovirus promoter or Rous sarcoma virus promoter;
• b. a DNA segment operably linked to said promoter;
• and
• c. sequences encoding a cis-ribozyme operably linked to said DNA segment,
• such that expression of the DNA segment produces a ribonucleotide sequence which is complementary to a ribonucleotide sequence transcribed from a gene, and said expressed ribonucleotide sequence is accumulated in the cell nucleus and inhibits the expression of said gene.

A method of inhibiting the expression of a gene in a cell, comprising introducing into a cell a DNA construct comprising:

• a. a human cytomegalovirus promoter or Rous sarcoma virus promoter;
• b. a DNA segment operably linked to said promoter;
• and
• c. sequences encoding a cis-ribozyme operably liked to said DNA segment,
• such that expression of the DNA segment produces a ribonucleotide sequence which is complementary to a ribonucleotide sequence transcribed from a gene, and said expressed ribonucleotide sequence is accumulated in the cell nucleus and inhibits the expression of said gene.

A method of inhibiting the expression of a gene in a cell, comprising the steps of:

• a. introducing into said cell a DNA construct comprising:
• i. a human cytomegalovirus promoter or a Rous sarcoma virus promoter;
• ii. a DNA segment operably linked to said transcriptional promoter;
• and
• iii. sequences encoding a cis-ribozyme operably linked to said DNA segment,
• such that expression of the DNA segment produces a ribonucleotide sequence which is complementary to a ribonucleotide sequence transcribed from a gene, and said expressed ribonucleotide sequence is accumulated in the cell nucleus and regulates the expression of said gene, whereby a transformed cell is obtained;
• and
• b. expressing the DNA segment to thereby inhibit the expression of said gene.