Autocatalytic replication of recombinant RNA
AssigneeThe Trustees of Columbia University in the City of New York
InventorsFred R. Kramer, Eleanor A. Miele, Donald R. Mills
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Abstract
This invention concerns recombinant RNA molecules comprising a recognition sequence for the binding of an RNA-directed RNA polymerase, a sequence for the initiation of product strand synthesis and a heterologous sequence of interest inserted at a specific site in the internal region of the recombinant molecule. Such recombinant RNA molecules are capable of serving as a template for the synthesis of complementary single-stranded molecules by RNA-directed RNA polymerase. The product molecules so formed are also capable of serving as a template for the synthesis of additional copies of the original recombinant RNA molecule. In a preferred embodiment of the invention Q.beta. replicase is used as the RNA-directed RNA polymerase.
Claims9
A recombinant single-stranded RNA molecule, wherein the sequence required for the initiation of product strand synthesis is a cytidine-rich 3' terminal sequence.
A recombinant, single-stranded RNA molecule capable of serving as a template in vitro for the synthesis of a complementary single-stranded RNA molecule by Q.beta. replicase which comprises MDV-1 RNA having the structure shown in FIG. 2 or a mutant thereof and a heterologous RNA sequence inserted into the MDV-1 RNA or the mutant thereof essentially without changing the secondary or tertiary structure of the MDV-1 RNA, the heterologous sequence of interest being inserted between two nucleotides, neither of the nucleotides being (a) hydrogen-bonded to any other nucleotide within the MDV-1 RNA, or (b) present within a Q.beta. replicase recognition sequence, or (c) present within a sequence for the initiation of product strand synthesis by Q.beta. replicase.
Dependencies are inferred from the claim text.