2'-O-alkyl oligoribonucleotides as antisense probes.

Iribarren, Adolfo M.; Sproat, Brian S.; Neuner, Philippe; Sulston, Ingrid; Ryder, Ursula; Lamond, Angus I.

doi:10.1073/pnas.87.19.7747

Cited by 129 publications

(59 citation statements)

References 8 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The sequences prepared (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17) are shown in Table 1. Average coupling yields for dimer incorporation were -90% as assessed by the trityl assay method.…”

Section: Resultsmentioning

confidence: 99%

“…Following chain assembly, the oligonucleotides were cleaved from the support, deprotected and purified as previously described (9). The single absorption band on polyacrylamide gel found for each oligomer indicated their high purity (>95%) and the retarded mobility of the sulfide containing oligomers (6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17) relative to the unmodified sequence (5) (9). How do we account for: (i) the general destabilizing effect of (3'-CH2-CH2-S-CH2-5') linkages, and (ii) for the discriminatory binding properties of sulfide strands containing 2'-substituted (OH and OMe) sugars?…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Structural basis for the RNA binding selectivity of oligonucleotide analogues containing alkylsulfide internucleoside linkages and 2′-substituted 3′-deoxyribonucleosides

et al. 1995

View full text Add to dashboard Cite

INTRODUCTIONIn this report we describe the synthesis of oligonucleotides containing sulfide-linked dinucleoside units, namely rT(2'OH)dT, rT(2'0Me)5dT, dT8rU(2'OMe) and dT(2o0Me)5rU(2oMe). We also describe the interactions of such oligomers with complementary DNA and RNA targets, and provide the structural basis for their remarkable RNA binding selectivity. In all cases, the Tm values of the SIP-chimera duplexes were lower than those of the corresponding unmodified duplexes. We attribute this to steric interactions between the 5' sulfur and the atoms of the nearby base/sugar residues. The 2'-substituents (i.e., 2'OH or 2'OMe) vicinal to the alkylsulfide internucleoside linkage significantly perturb the structure and stability of the duplexes formed with DNA, and more so than with RNA. The introduction of three rT(2'OH)sdTp ( ' (5,6).Our research in the antisense field has focused on oligonucleotide analogues in which some of the phosphodiester groups are replaced by the dialkyl sulfide linkage 3'-CH2-CH2-S-CH2-5' (7-11). This linkage is non-hydrolyzable, non-ionic, and its length approximates that of a phosphodiester linkage reasonably well. More recently, we focused attention on the preparation of rT(20OH)SdT, rT(2 OMe)SdT, dTsrU(2'OMe) and rT(2'OMe)SrU(2'OMe) dimer units as well as their 3'-O-phosphoramidite derivatives for incorporation into oligonucleotides (11,12 (55°C, 16 h). The ammonia solution was collected by centrifuging the Eppendorf tube and then run through a Sephadex NAP-10 column which was pre-washed with 15 ml deionized water. The first eluting fractions (1.5 ml) were lyophilized to dryness and the resulting oligomer was redissolved in 1 ml of water. Purity of each oligomer was confmed by PAGE (26% polyacrylamide-7 M urea) run at 4°C, at a current of 10 mA, for 6-8 h (9). Melting experimentsMelting curves were measured in 10 mM NaH2PO4/Na2HPO4, 1 M NaCl buffer (pH 7.0) and analyzed the same way as described in reference 9. Circular dichroismThe CD spectrum ofeach sample was measured on a Jasco 500-C spectropolarimeter with a Jasco DP-J800/300 data processor. The cell temperature was kept at 50C. The concentration of the oligonucleotides were -2.5 jM (each strand).

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Structural basis for the RNA binding selectivity of oligonucleotide analogues containing alkylsulfide internucleoside linkages and 2′-substituted 3′-deoxyribonucleosides

et al. 1995

View full text Add to dashboard Cite

show abstract

“…Equal amounts (2-4 g) of poly(A) RNA from both reference and experimental samples were used as templates to synthesize fluorescently labeled cDNA probes for hybridization to the microarrays. The RNA was reverse transcribed in the presence of 5-(3-aminoallyl)-2Ј-deoxyuridine 5Ј triphosphate (18). The reference sample was coupled to fluorescent dye (Cy)3, and the experimental sample was coupled to Cy5.…”

mentioning

confidence: 99%

Genome-wide study of aging and oxidative stress response inDrosophilamelanogaster

Zou

Meadows²,

Sharp³

et al. 2000

Proc. Natl. Acad. Sci. U.S.A.

287

173

View full text Add to dashboard Cite

Aging is a universal but poorly understood biological process. Free radicals accumulate with age and have been proposed to be a major cause of aging. We measured genome-wide changes in transcript levels as a function of age in Drosophila melanogaster and compared these changes with those caused by paraquat, a free-radical generator. A number of genes exhibited changes in transcript levels with both age and paraquat treatment. We also found genes whose transcript levels changed with age but not with paraquat treatment. This study suggests that free radicals play an important role in regulating transcript levels in aging but that they are not the only factors. This genome-wide survey also identifies candidates for molecular markers of aging.A ging is a biological process universal to eukaryotic organisms, and its underlying mechanisms are under intensive study. Genetic analyses of yeast, nematode, fly, and mouse have uncovered a number of genes, whether mutated or misexpressed, that would increase the lifespans of these organisms (1). These genes include superoxide dismutase, a free-radical scavenger; methuselah, a potential G protein-coupled receptor, in Drosophila melanogaster; and p66 shc , an oxidative stress-response gene, in mice (2-5). Lifespan-related genes in Caenorhabditis elegans include clk-1, a gene involved in ubiquinone biosynthesis, and a group of genes involved in an insulin receptor-like signaling pathway: daf-2, age-1, and daf-16 (6, 7). Many mutations that increase lifespan also confer resistance to oxidative stress (1). This finding supports the free-radical hypothesis of aging, which suggests that reactive oxygen species that accumulate with increasing age cause oxidative damage to macromolecules (including nucleic acids, proteins, and lipids) and are causally linked to aging and death (8, 9). Free radicals have been found to regulate the expression of a number of genes that include antioxidant defense genes involved in repairing oxidative damage, as well as genes involved in inducing apoptosis (10, 11). The aging process is also accompanied by changes in the expression patterns of a number of genes (12)(13)(14). How the regulation of gene expression in aging correlates with that in response to oxidative stress, however, is understood poorly. Materials and MethodsConstruction of Microarray. We constructed a microarray containing 7,829 expressed sequence tags (ESTs). This set includes 221 ESTs provided by our lab and 7,608 ESTs generously supplied by K. White (Stanford University, Stanford, CA) and K. Burtis (University of California, Davis). The ESTs were amplified (as described by White et al., ref. 15), and the DNA was mechanically spotted onto polylysine-coated slides (as described by DeRisi et al., ref. 16).Calculation of Survival Rate and Preparation of Fly Tissues. Male flies (w 1118 ) raised in standard cornmeal agar medium, were collected within 24 h after eclosion (17). Approximately 200 flies were maintained in constant darkness in each food bottle at 26-27°C and 60-70% humidity an...

show abstract

“…Results from a number of laboratories have shown that oligonucleotides which contain 2'-0-alkyl substituents, such as 2'-0-methylribose, form very stable duplexes with complementary RNA target molecules (12)(13)(14)(15)(16). The improved binding constants observed with oligo-2'-O-methylribonucleotides and the enhanced nuclease resistance of the phosphorothioate or methylphosphonate oligomers could be used to advantage to design effective antisense oligonucleotides.…”

Section: Introductionmentioning

confidence: 99%

Interactions of oligonucleotide analogs containing methylphosphonate internucleotide linkages and 2′-O-methylribonucleosides

et al. 1994

View full text Add to dashboard Cite

show abstract

2'-O-alkyl oligoribonucleotides as antisense probes.

Cited by 129 publications

References 8 publications

Structural basis for the RNA binding selectivity of oligonucleotide analogues containing alkylsulfide internucleoside linkages and 2′-substituted 3′-deoxyribonucleosides

Structural basis for the RNA binding selectivity of oligonucleotide analogues containing alkylsulfide internucleoside linkages and 2′-substituted 3′-deoxyribonucleosides

Genome-wide study of aging and oxidative stress response inDrosophilamelanogaster

Interactions of oligonucleotide analogs containing methylphosphonate internucleotide linkages and 2′-O-methylribonucleosides

Contact Info

Product

Resources

About