Improved probes using nucleosides containing 3-dezauracil analogs
AssigneeMonsanto Company
InventorsThomas E. Rogers, Steven H. Gray, Balekudru Devadas, Steven P. Adams
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Abstract
The deazauracil containing probes of the invention are able to withstand higher temperatures, thereby allowing unmatched probes and mismatched probes to be washed off at higher hybridization stringency, thereby eliminating background readings and improving ease and accuracy of probe use.
Claims25
A single-stranded nucleic acid probe which comprises a nucleoside residue of the formula:
- ##STR76## in which R.sub.1 and R.sub.2 independently are C.sub.1 -C.sub.5 alkyl, C.sub.2 -C.sub.5 alkenyl, halo or hydrogen, R.sub.3 is hydrogen, hydroxy, C.sub.6-14 aryloxy or C.sub.1 -C.sub.5 alkoxy and in which the nucleoside residue is at least five nucleosides from both ends of the probe and said probe is capable of hybridizing with a target nucleic acid and said hybridizing is mediated by the presence of nondeazauracil nucleotide which are contained within said probe residues.
The probe which is selected from the group consisting of GGCAGCTTYCACATAGTCCA, AGCTTCCAYATGGTCC, AGCTTCCAYATGGTCCA, GCAGCTTYCACATGGTCC, GGCAGCTTYCACATGGTCCA, ATGAATCYGAACAATCG, ATGAATCCGAAYAATCG, TAGCGGCGACGYACGTACAC, CTCTAGGYTGTCGGCCAGCC, TAGGCAGCTTCCAYCTGGTCCAGTAGC, and GGCAGCTTCCACATGGTCYA in which Y=4-hydroxy-1(2'-deoxy-beta-D-erythro-pentofuranosyl)-2(1H)-pyridone.
The probe having the following sequence:
- GGCAGCTTXCACATGGTCCA, in which X=3-iodo-4-hydroxy-1-(2'-deoxy-beta-D-erythro-pentofuranosyl)-2(1H)-pyrido ne.
The probe having the following sequence:
- GGCAGCTTXCACATGGTCCA, in which X=5-iodo-4-hydroxy-1-(2'-deoxy-beta-D-erythro-pentofuranosyl)-2(1H)-pyrido ne.
The probe having the following sequence:
- GGCAGCTTXCACATGGTCCA, in which X=4-0-diphenylcarbamoyl-3-ethenyl-1-(2'-1-deoxy-beta-D-erythropentofuranos yl)-2(1H)-pyridone.
A method for preparing hybrid DNA or RNA molecules which comprises hybridizing a target nucleic acid molecule to a single-stranded nucleic acid probe which comprises a nucleoside residue of the formula:
- ##STR77## in which R.sub.1 and R.sub.2 independently are C.sub.1 -C.sub.5 alkyl, C.sub.2 -C.sub.5 alkenyl, halo or hydrogen, R.sub.3 is hydrogen, hydroxy, C.sub.6 -C.sub.14 aryloxy or C.sub.1 -C.sub.5 alkoxy, and in which the nucleoside residue is at least five nucleosides from both ends of the probe and said probe undergoes hybridization with a target nucleic acid and said hybridization is mediated by the presence of non-deazauracil nucleoside residues which are contained within said probe.
Dependencies are inferred from the claim text.