Modification of RNA, producing an increased transcript stability and translation efficiency
It was the object of the present invention to provide RNA with increased stability and translation efficiency and means for obtaining such RNA. It should be possible to obtain increased grades of expression by using said RNA in gene therapy approaches.
1. An RNA molecule having a nucleic acid sequence that is complementary to a template nucleic acid molecule comprising:
(a) a first nucleic acid sequence having at least two copies of a 3′-untranslated region of a human beta-globin gene, wherein each copy of the 3′-untranslated region of the human beta-globin gene comprises a region of SEQ ID NO: 1 that extends from the termination codon to the poly(A) attachment signal; and
(b) a second nucleic acid sequence comprising a transcribable nucleic acid sequence or a nucleic acid sequence for introducing the transcribable nucleic acid sequence wherein the transcribable nucleic acid sequence comprises a nucleic acid sequence coding for a peptide or protein;
wherein the nucleic acid sequence of the RNA molecule is complementary over the entire length of the first and second nucleic acid sequences to the template nucleic acid molecule.
2. The RNA molecule of claim 1 , further comprising a 3′ end having at least 20 consecutive A nucleotides, at least 40 consecutive A nucleotides, at least 80 consecutive A nucleotides, at least 100 consecutive A nucleotides, or about 120 consecutive A nucleotides.
3. A method comprising: transecting a host cell with the RNA molecule of claim 1 .
4. The method claim 3 , wherein the host cell is an antigen-presenting cell.
5. The method of claim 3 , wherein the host cell is a dendritic cell, a monocyte or a macrophage.
6. The RNA molecule of claim 1 wherein the first nucleic acid sequence is 3′ to the second nucleic acid sequence.
7. An RNA molecule having a nucleic acid sequence comprising:
(a) a first nucleic acid sequence having at least two copies of a 3′-untranslated region of a human beta-globin gene, wherein each copy of the 3′-untranslated region of the human beta- globin gene comprises a region of SEQ ID NO: 1 that extends from the termination codon to the poly(A) attachment signal; and
(b) a second nucleic acid sequence comprising a nucleic acid sequence coding for a peptide or protein, wherein the second nucleic acid sequence is 5′ to the first nucleic acid sequence and can be transcribed to give a common transcript.
8. The RNA molecule of claim 7 , further comprising a 3′ end having at least 20 consecutive A nucleotides, at least 40 consecutive A nucleotides, at least 80 consecutive A nucleotides, at least 100 consecutive A nucleotides, or about 120 consecutive A nucleotides.
9. A method comprising: transfecting a host cell with the RNA molecule of claim 7 .
10. The method of claim 9 , wherein the host cell is an antigen-presenting cell.
11. The method of claim 9 , wherein the host cell is a dendritic cell, a monocyte or a macrophage.