The Equal Employment Opportunity Commission and people with mental retardation.
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Commission on Professional and Hospital Activities↗
Biomedical subjects
P S Miller
Publications and source records attributed to P S Miller.
Effects of the reproductive cycle and age on calcium and phosphorus metabolism and bone integrity of sows.
The purpose of this study was to determine the effects of stage of the reproductive cycle and age on Ca and P metabolism and bone integrity of sows. Five-day balance studies were conducted with first- and fifth-parity sows, and sows were slaughtered during the last trimester of gestation, at the end of lactation, or during the last trimester of the subsequent gestation. First-parity sows were studied during their first gestation (n = 11), first lactation (n = 10), or second gestation (n = 10). Fifth-parity sows were examined during their fifth gestation (n = 10), fifth lactation (n = 9), or sixth gestation (n = 9). All sows were fed 1.9 kg/d of a common diet (.76% Ca and .63% P) during gestation and were allowed ad libitum access to the same diet during lactation. Digestibilities of Ca and P were much greater during lactation than during gestation. During gestation, young sows absorbed and retained more Ca and P than did mature sows. However, during lactation, mature sows consumed more feed, and therefore Ca and P, and retained more Ca and P than did young sows. Bones of mature sows were larger, more mineralized, and stronger than the bones of young sows. Bone weight and strength decreased during lactation and increased during the subsequent gestation. Changes in weight and strength were greater in young sows than in mature sows.
Aging↗
Influence of sample orientation on prediction of fresh ham lean content by electromagnetic scanning.
To evaluate the effect of orientation of hams during electromagnetic scanning on the estimation of dissected lean content, hams were scanned horizontally, posterior first (POS) or dorsal first (DOR), and vertically, medial side (aitchbone) first (MED; standing on the butt face). Weight and percentage of dissected lean were estimated using scan peak for each orientation, ham weight, and fat thickness. The mean scan peak for the MED orientation was approximately twice as great as peaks for the POS and DOR orientations, which suggests that this orientation may offer greater predictive accuracy by reducing the signal to noise ratio. Results, however, indicated that all orientations were equally effective at predicting lean weight and percentage, with R2 values of .95 and .75 and root mean square errors of .21 kg and 2.6%, respectively.
Animals↗
Triplex formation by oligonucleotides containing novel deoxycytidine derivatives.
Homopurine sequences of duplex DNA are binding sites for triplex-forming oligodeoxyribopyrimidines. The interactions of synthetic duplex DNA targets with an oligodeoxyribopyrimidine containing N4-(6-amino-2-pyridinyl)deoxycytidine (1), a nucleoside designed to interact with a single C-G base pair interruption of the purine target tract, was studied by UV melting, circular dichroism spectroscopy and dimethylsulfate alkylation experiments. Nucleoside 1 supports stable triplex formation at pH 7.0 with formation of a 1-Y-Z triad, where Y-Z is a base pair in the homopurine tract of the target. Selective interaction was observed when Y-Z was C-G, although A-T and, to a lesser extent, T-A and G-C base pairs were also recognized. The circular dichroism spectra of the triplex having a 1-C-G triad were similar to those of a triplex having a C(+)-G-C triad, suggesting that the overall structures of the two triplexes are quite similar. Removal of the 6-amino group from 1 essentially eliminated triplex formation. Reaction of a triplex having the 1-C-G triad with dimethylsulfate resulted in a 50% reduction of methylation of the G residue of this triad. In contrast, the G of a similar triplex containing a U-C-G triad was not protected from methylation by dimethylsulfate. These results are consistent with a binding mode in which the 6-amino-2-pyridinyl group of 1 spans the major groove of the target duplex at the 1-C-G binding site and forms a hydrogen bond with the O6 of G. An additional stabilizing hydrogen bond could form between the N4 of the imino tautomer of 1 and the N4 amino group of C.
Alkylation↗
Triplex formation by a psoralen-conjugated oligodeoxyribonucleotide containing the base analog 8-oxo-adenine.
Oligodeoxyribonucleotides containing thymidine and 8-oxo-2'-deoxyadenosine can form pyr.pur.pyr type triplexes with double-stranded DNA. Unlike triplexes whose third strands contain thymidine and deoxycytidine, the stability of these triplexes is independent of pH. We have prepared d-ps-TAAATAAATTTTTAT-L [I(A)], where A is 8-oxo-2'-deoxyadenosine, ps is 4'-hydroxymethyl-4,5',8- trimethylpsoralen and L is a 6-amino-2-(hydroxymethyl)hexyl linker. The oligomer is designed to interact with a homopurine sequence in the promoter region of the human gene coding for the 92 kDa form of collagenase type IV. Oligomer I(A) and oligomer I(C), which contains 2'-deoxycytidine in place of 8-oxo-2'-deoxycytidine, both form stable triplexes at pH 6.2, but only I(A) forms a stable triplex with a model duplex DNA target at pH 7.5, as determined by UV melting experiments. Triplex formation is stabilized by the presence of the psoralen group. Upon irradiation both I(A) and I(C) form photoadducts with the DNA target at pH 6.2, but only I(A) forms a photoadduct at pH 7.5. In these photoreactions oligomer I(A) appears to selectively form a photoadduct with a C in the purine-rich strand of the duplex target. Although a T residue is present in the pyrimidine-rich strand of the target at the duplex/triplex junction, essentially no adduct formation takes place with this strand, nor is interstrand cross-linking observed. The extent of photoadduct formation decreases with increasing temperature, behavior which is consistent with the UV melting curve of the triplex. A tetramethylrhodamine derivative of I(A) was prepared and found to cross-link less extensively than I(A) itself. Oligomer I(A) is completely resistant to hydrolysis when incubated for 24h in the presence of 10% fetal bovine serum at 37 degree C, although it is hydrolyzed by S1 nuclease. The properties of oligomer I(A) suggest that 8-oxo- containing oligomers may find utility as antigene oligonucleotide reagents.
Adenine↗
Development of antisense and antigene oligonucleotide analogs.
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Animals↗
Inhibition of human collagenase activity by antisense oligonucleoside methylphosphonates.
Oligodeoxyribonucleoside methylphosphonates (d-OMP) were synthesized whose sequences are complementary to sequences found in the mRNA coding for the 72-kDa (MMP-2) or 92-kDA (MMP-9) forms of human collagenase i.v., matrix metalloproteinases (MMP) whose excessive secretion correlates with the metastatic potential of tumor cells. The effects of these oligomers on MMP-2 and MMP-9 activities secreted by HT1080 cells, a human fibrosarcoma cell line, were studied using a gelatin zymography assay. A d-OMP, M2.3, complementary to nucleotides 14 to 28 of the initiation codon region of MMP-2 mRNA selectively inhibited MMP-2 activity, whereas a d-OMP, M9.1, which was targeted to nucleotides -19 to -5 of the 5'-untranslated region of MMP-9 mRNA selectively inhibited MMP-9 activity over the concentration range 5-50 microM. At 100 microM concentration, both M2.3 and M9.1 inhibited the activities of both MMP-2 and MMP-9. These oligomers were completely stable under cell culture conditions and did not appear to adversely affect cell growth after 48 hours at concentrations up to 100 microM, although 100 microM M9.1 did reduce cell growth 30% after prolonged, 120-hours exposure. Other d-OMP tested either had no effect on collagenase activity or inhibited both MMP-2 and MMP-9 activities. The latter oligomer was complementary to MMP-2 mRNA and partially complementary to MMP-9 mRNA. Oligomer M2.3 was also tested for its effects on the morphology of malignant human lung cells, BZR-T33, growing on the surface of reconstituted base membrane, Matrigel, in culture. In absence of oligomer, the BZR-T33 cells formed extensive networks indicative of the ability of the cells to invade the Matrigel substrate. In the presence of 100 microM M2.3, BZR-T33 formed colonies of rounded cells, a morphology typical of noninvasive cells. Other non-complementary d-OMP had no effect on the morphology of BZR-T33 under these conditions. These results suggest that antisense d-OMP may be useful for inhibiting expression of collagenase in human tumor cells and for studying the role of collagenase expression in tumor cell metastasis.
Collagenases↗
Response of pigs to space allocation and diets varying in nutrient density.
Three experiments were conducted to determine the main and interacting effects on growth performance of floor space allowance and dietary lysine and energy concentrations for growing-finishing pigs. In each experiment, space allocations of .56 or .78 m2/pig were achieved with 14 or 10 pigs per pen, respectively. In Exp. 1, diets investigated were National Research Council (NRC) recommended nutrient densities, NRC plus 5% added fat (F), NRC plus .15% added L-lysine. HCl (L), and NRC plus fat plus lysine (FL) in a 4 x 2 x 2 factorial arrangement of diets with space and season (winter vs summer). In Exp. 2 and 3, a 2 x 3 factorial treatment arrangement was used to investigate space and diet effects. Diets investigated contained 0, 2.5, or 5% added fat (choice white grease [Exp. 2]; tallow [Exp. 3]) with L-lysine.HCl added to maintain a constant lysine:ME ratio. In Exp.1, pigs given .56 m2/pig vs .78 m2/pig ate less feed (P < .001) and grew slower (P < .001) with no difference in gain:feed ratio or carcass lean percentage. Feed intake was decreased (P < .005) and gain:feed ratio increased (P < .001) for the FL vs L diet. In Exp. 2, pigs given .56 m2/pig vs .78 m2/pig grew slower (P < .001) with a poorer gain:feed ratio (P < .05) and a slower rate of lean gain (P < .05). In Exp. 3, pigs given .56 m2/pig vs .78 m2/pig grew slower (P < .05) with no difference in gain:feed ratio and a slower rate of lean gain (P < .005). There was a linear improvement in ADG (P < .01, Exp. 2; P = .011, Exp. 3) and gain:feed ratio (P < .001, Exp. 2 and 3) with increasing amounts of fat in the diet. The lack of space x diet interactions in these experiments suggests that the reduction in ADG associated with the reduction in ADFI for pigs given less space is independent of dietary lysine and energy concentrations.
Animal Nutritional Physiological Phenomena↗
Disturbances in the soil: finding buried bodies and other evidence using ground penetrating radar.
Ground penetrating radar (GPR) is an efficient and effective means to search for buried evidence, whether it be a clandestine grave, formal burial, or certain missing articles from a crime scene. The procedures for GPR used by the U.S. Army Central Identification Laboratory, Hawaii (CILHI), are the result of several years of experimentation on a variety of ground surfaces in Hawaii, Southeast Asia and the mainland U.S. This remote sensing method does not usually provide direct information that there is a body or other specific object beneath the ground. Most of the time the GPR has been used to determine where a target object is not located. The key feature of GPR is that it can detect recent changes in shallow soil conditions caused by the disturbance of soil and the intrusion of different material. Using the methods described here, the investigator should be able to determine the precise metric grid coordinates for a subsurface disturbance, as well as the approximate size, the general shape, and the depth of the buried material. Success will vary with soil conditions. The conditions suitable or not practical for using GPR are summarized. This remote sensing technology can have wider use in crime scene investigations due to the recent introduction of more user-friendly software and more portable hardware.
Burial↗
Inhibition of herpes simplex virus replication by antisense oligo-2'-O-methylribonucleoside methylphosphonates.
Antisense oligonucleoside methylphosphonates complementary to the 12 nucleotides found at the intron/exon junction of the splice acceptor site of herpes simplex virus type 1 (HSV-1) immediate early mRNAs 4 and 5 were synthesized. The methylphosphonate oligomers contained either 2'-deoxyribose nucleosides, d-OMPs, or 2'O-methylribose nucleosides, mr-OMPs. At 37 degrees C, the affinity of the mr-OMP for a complementary 12-mer RNA target was approximately four times higher than that of the corresponding d-OMP as measured by a constant activity gel electrophoresis mobility shift assay. An mr-OMP whose sequence contained two mismatched bases did not bind to the RNA target under these conditions. The mr-OMP also showed improved ability to inhibit HSV-1 replication in HSV-1 infected Vero cells in culture. Thus the IC50 of the mr-OMP was five times less than that of the d-OMP. No inhibition was observed by the mismatched mr-OMP, and no inhibition of herpes simplex virus type 2 (HSV-2) replication was observed with any of the oligomers. These results demonstrate a direct correlation between oligomer binding affinity and antisense activity in cell culture and suggest that oligo-2'-O-methylribonucleoside methylphosphonates are promising candidates for development of effective antisense reagents.
Animals↗
The effects of dietary protein concentration on compensatory growth in barrows and gilts.
An experiment using 120 crossbred pigs (60 barrows and 60 gilts) was conducted to determine the effect of dietary protein concentration on the compensatory response in growth after a period of feed restriction. Thirty pigs were allowed ad libitum (AL) access to one of five corn-soybean meal diets (13.1 to 18.4% CP) from approximately 42 to 102 kg BW. Thirty additional pigs were restricted (R) to a maintenance amount of a 14.4% CP diet for 21 d, after which they were allotted to one of the five diets and allowed ad libitum access to feed until they reached 102 kg. Restricted pigs had lower ADFI, ADG, and ADG/ADFI (P < .05) than did AL pigs. During the postrestriction period, R pigs had higher ADFI and ADG (P < .001) and tended to be more efficient (P < .08) than AL pigs. Average backfat thickness was not different (P > .8) between AL and R pigs. Lean percentage (5% fat) was not affected (P > .5) by feeding regimen but increased linearly (P < .1) as protein level increased. Organ weights (expressed as a percentage of empty BW.75) were similar in AL and R pigs. However, kidney and stomach weights differed among protein levels (P < .01), as did liver weight (P < .06), with a linear increase (P < .02) in both kidney and liver weights to dietary protein level. Barrows had higher ADFI and ADG and greater average backfat thickness (P < .02) than gilts. Barrows had smaller LMA, a lower percentage of lean (P < .001), and higher average backfat (P < .005) than gilts. Liver weight was heavier (P < .02) and small intestine weight tended to be heavier (P < .07) for barrows than for gilts. Kidney, spleen, and lungs were heavier (P < .05) in gilts than in barrows. Livers from barrows tended to contain more protein (P < .06) and less fat (P < .05) than livers from gilts. These results indicate that a compensatory growth response occurred during relimentation, and that the effects of dietary protein concentration on growth rate and carcass measurements were similar in both AL and R pigs.
Animals↗
Pork characteristics as affected by two populations of swine and six crude protein levels.
Previous research has investigated the effect of genetic line and dietary CP on gilt growth and carcass composition. The aim of this research was to characterize lean tissue composition, color, tenderness, and water-holding capacity as affected by protein level and genetic lines. Thirty-six Gene Pool pigs, a population with low lean growth potential, and 36 Hampshire pigs, a population with a high lean growth potential, had ad libitum access to diets consisting of 10, 13, 16, 19, 22, or 25% CP. Longissimus muscles of Hampshire pigs were lighter, more red, and more yellow than those from Gene Pool pigs (P< .01). Redness and yellowness of longissimus muscles decreased linearly (P < .01) as protein level increased. Chops from Hampshire pigs required less force and less total energy to shear but had lower cooking yields and water-holding capacity than chops from Gene Pool pigs (P < .01). Chops from pigs fed 19 and 22% protein required greater force (quadratic, P < .01) and more total energy (quadratic, P < .05) to shear. Objective color differences revealed that restructured, cured hams from Hampshire pigs were lighter, less red, and had less intense cured color (P < .01). The Hampshire line of gilts produced pork muscle that was leaner, more pale in color, more tender, and lower in water-holding capacity than the Gene Pool line. Level of dietary CP caused alterations in tenderness and lean composition, especially when diets contained less protein.
Animals↗
Changes in plasma urea concentration can be used to determine protein requirements of two populations of pigs with different protein accretion rates.
The experiment had two objectives: 1) to determine the protein requirements of two strains of growing-finishing pigs based on growth performance, carcass characteristics, tissue accretion rates, and organ weights and 2) to evaluate whether protein requirements can be determined from changes in plasma urea concentration. Forty-six Gene Pool (GP) and 46 Hampshire (H) gilts with an initial BW of 28.5 kg were used. Pigs were allotted to two trials of a randomized complete block experiment with a 2 x 6 factorial arrangement of treatments. Five pigs from each strain were randomly selected and slaughtered at the beginning of each of the two trials. The remaining 72 pigs were individually penned and allotted to one of six dietary treatments (10, 13, 16, 19, 22, or 25% CP). Pigs remained on the experiment until the mean weight of a treatment group within each strain reached 115 kg (16 wk for GP and 14 wk for H), at which time all pigs of that strain were slaughtered. The only strain x protein level interactions that were detected were for carcass protein and water accretion rates. Gene Pool pigs grew less rapidly and utilized feed less efficiently than H pigs (P < .001). Average daily gain (quadratic, P < .05) and ADG/ADFI (quadratic, P < .05) were increased as protein level increased until a plateau was reached. Backfat depths were decreased (linear, P < .001) and longissimus muscle areas were increased (linear, P < .001) as protein level increased. Protein accretion rate was lower (P < .01) and fat accretion was higher (P < .01) in GP pigs than in H pigs. Protein accretion increased (quadratic, P < .001) and fat accretion decreased (linear, P < .001) with increasing dietary protein level. Examination of the response of plasma urea concentration over time suggested that GP pigs required 13% CP from 30 to 80 kg and 10% CP thereafter, whereas H pigs required 19% CP from 30 to 45 kg, 16% CP from 45 to 100 kg, and 13% CP thereafter.
Analysis of Variance↗
Interactions of oligonucleotide analogs containing methylphosphonate internucleotide linkages and 2'-O-methylribonucleosides.
The interactions of oligonucleotide analogs, 12-mers, which contain deoxyribo- or 2'-O-methylribose sugars and methylphosphonate internucleotide linkages with complementary 12-mer DNA and RNA targets and the effect of chirality of the methylphosphonate linkage on oligomer-target interactions was studied. Oligomers containing a single Rp or Sp methylphosphonate linkage (type 1) or oligomers containing a single phosphodiester linkage at the 5'-end followed by 10 contiguous methylphosphonate linkages of random chirality (type 2) were prepared. The deoxyribo- and 2'-O-methylribo- type 1 12-mers formed stable duplexes with both the RNA and DNA as determined by UV melting experiments. The melting temperatures, Tms, of the 2'-O-methylribo-12-mer/RNA duplexes (49-53 degrees C) were higher than those of the deoxyribo-12mer/RNA duplexes (31-36 degrees C). The Tms of the duplexes formed by the Rp isomers of these oligomers were approximately 3-5 degrees C higher than those formed by the corresponding Sp isomers. The deoxyribo type 2 12-mer formed a stable duplex, Tm 34 degrees C, with the DNA target and a much less stable duplex with the RNA target, Tm < 5 degrees C. In contrast, the 2'-O-methylribo type 2 12-mer formed a stable duplex with the RNA target, Tm 20 degrees C, and a duplex of lower stability with the DNA target, Tm < 5 degrees C. These results show that the previously observed greater stability of oligo-2'-O-methylribonucleotide/RNA duplexes versus oligodeoxyribonucleotide/RNA duplexes extends to oligomers containing methylphosphonate linkages and that the configuration of the methylphosphonate linkage strongly influences the stability of the duplexes.
Adenosine↗
Effect of target structure on cross-linking by psoralen-derivatized oligonucleoside methylphosphonates.
A series of psoralen-derivatized oligodeoxyribonucleoside methylphosphonates were examined for their abilities to cross-link to DNA and RNA oligonucleotide targets. These targets were designed to have either a random coil or a hairpin structure in solution. The methylphosphonate oligomers cross-linked with approximately the same rates to the random coil DNA and RNA targets, although the extent of cross-linking to the DNA target was higher than that to the RNA target. For a given methylphosphonate sequence, cross-linking decreased as the temperature increased, and this behavior paralleled the interaction of the oligomer with the target as determined by ultraviolet melting experiments. The oligomers also cross-linked efficiently with the DNA hairpin target, but little or no cross-linking was observed with the RNA hairpin. In the case of these hairpin targets, the extent of cross-linking was dependent upon the location of the oligomer binding site relative to the stem and loop regions of the hairpin. The lack of reactivity with the RNA hairpin may be due to the high stability of the stem of this target versus that in the DNA target and the relatively lower efficiency of binding of the methylphosphonates to RNA versus DNA targets. The sequences of the oligomers are complementary to vesicular stomatitis virus M-protein mRNA. One of the oligomers was tested, and was found to cross-link at 20 degrees C to VSV N-mRNA to approximately the same extent as observed for cross-linking with the random coil RNA target, suggesting that the mRNA binding site for the oligomer most likely is in a somewhat open conformation.(ABSTRACT TRUNCATED AT 250 WORDS)
Base Sequence↗
Rare alleles, MHC and captive breeding.
In recent years, more detailed genetic information has become available for individuals of endangered species in captive breeding programs. There have been suggestions that this information be used to identify rare alleles, particularly those at the MHC, that can be subsequently selected for captive breeding programs. First, we summarize the current information on the MHC relevant to conservation genetics, so that such a possible breeding program is seen in a proper perspective. For example, very few specific alleles at the MHC have been identified as selectively advantageous, even though there has been substantial effort to find such alleles in humans and a few other organisms. Further, many of the balancing selection models suggested for MHC variation are based on heterozygotes in general having a higher fitness than homozygotes and not on specific selectively advantageous alleles. Because there is no detailed data on MHC variability in captive populations, we used transferrin data in Przewalski's horses to evaluate a breeding program to select for rare alleles. In this species, one individual, 1060, has been identified to have the transferrin allele J. We determine the effect on founder contribution of multiply mating 1060 to increase the number of copies of this allele. Since there were 485 individuals in the population at this time, this extra mating had little detrimental effect on the distribution of founder contributions and the number of founder equivalents. We then selected 65, an ancestor of 1060, which had a high likelihood of being the individual that passed on the J allele in the lineage of 1060. We examined the effect of increasing the number of copies of alleles of 65 at a time when the population had only 22 other individuals. In this case, even though the founder contributions were changed more, there was also little effect on the founder contributions and the number of founder equivalents. Overall, it appears that selection that results in a limited change in the number of copies of rare alleles may not always have an overall detrimental effect. However, because other pedigrees may have very different properties, it is essential to perform a detailed pedigree analysis of any such selective breeding program to determine its effect before such a selection program is implemented.
Alleles↗
Interactions of psoralen-derivatized oligodeoxyribonucleoside methylphosphonates with vesicular stomatitis virus messenger RNA.
The ability of oligonucleotides to interact selectively with their targets is an important consideration in the design of antisense oligonucleotides. This is especially important in the case of antisense oligomers, such as psoralen-derivatized oligomers, which can irreversibly bind to their targets. We have studied the interactions of a series of psoralen-derivatized antisense oligonucleoside methylphosphonates with the mRNAs of vesicular stomatitis virus (VSV), mRNAs that have a high degree of sequence homology. Cross-linking reactions were carried out under conditions of low ionic strength in order to reduce mRNA secondary structure. A 12-mer, whose sequence was complementary to VSV M-mRNA and partially complementary to sequences found in N, NS, and G mRNA cross-linked extensively to N-message. On the other hand, 16-mers whose sequences were uniquely complementary to binding sites on N- or M-mRNA specifically and efficiently cross-linked to their targeted mRNAs over the temperature range 0 degree to 37 degrees C. A reverse transcriptase-catalyzed primer extension assay was used to show that one of the N-specific oligomers cross-linked at the expected site on N-mRNA and to estimate the extent of cross-linking. The results demonstrate that psoralen-derivatized oligonucleoside methylphosphonates can cross-link in a sequence-specific manner if the sequences of these oligomers are chosen carefully so as to avoid extensive partial complementarity with other mRNA sequences.
Animals↗
Properties of exonuclease-resistant, psoralen-conjugated oligodeoxyribonucleotides in vitro and in cell culture.
We have prepared oligodeoxyribonucleotides that are modified at the 3'-terminal with N4-(4-aminobutyl)deoxycytidine and derivatized at the 5'-end with a 4'-([N-(aminoethyl)amino]methyl)-4,5',8-trimethylpsoralen, (ae)AMT, and whose sequences are complementary to vesicular stomatitis virus (VSV), N-protein mRNA, (ae)AMT-II, or VSV M-protein mRNA, (ae)AMT-III. (ae)AMT-II cross-links exclusively to VSV N-mRNA when a mixture of the oligomer and poly(A+) RNA from VSV-infected cells is irradiated in vitro with long wavelength UV light at either 20 degrees or 37 degrees C. N4-(4-Aminobutyl)deoxycytidine at the 3'-end of (ae)AMT-II does not appear to affect the binding or cross-linking of the oligomer to its target RNA. Oligomer (ae)AMT-II is completely resistant to hydrolysis by the 3'-5'-exonuclease activity found in fetal calf serum whereas a similar oligomer, (ae)AMT-I, which contains a 3'-terminal deoxycytidine, is hydrolyzed within 30 min when incubated at 37 degrees C. Intact (ae)AMT-II was found in both the cell lysate and cell culture medium after 12 hr of incubation with mouse L-cells along with d-(ae)AMTpT, which appears to result from endonuclease degradation of the oligomer. In contrast no intact (ae)AMT-I was found in either the cell lysate or the culture medium after 1 hr incubation. Although 10 microM (ae)AMT-II had no effect on VSV-protein synthesis in either unirradiated or UV-irradiated VSV-infected mouse L-cells, 10 microM (ae)AMT-III inhibited VSV protein synthesis 30% in irradiated cells. These results show that introduction of a N4-(4-aminobutyl)deoxycytidine at the 3'-end of an oligodeoxyribonucleotide significantly increases the resistance of the oligomer to degradation by 3'-5'-exonucleases but does not interfere with its ability to bind selectively to complementary RNA. Further derivatization with psoralen creates an oligomer that can be triggered to cross-link with RNA in a sequence-specific manner, is taken up intact by mammalian cells in culture, and exhibits biological activity. In combination, these two modifications endow the oligodeoxyribonucleotide with novel properties that could be exploited in the design of antisense or antigene reagents for use in controlling gene expression in mammalian cells.
Animals↗