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PubMed1989

Activation and repression of a beta-globin gene in cell hybrids is accompanied by a shift in its temporal replication.

To investigate whether a switch in the transcriptional activity of a gene is associated with a change in the timing of replication during the S phase, we examined the replication timing of the beta-globin genes in two different types of somatic cell hybrids. In mouse hepatoma (Hepa 1a) x mouse erythroleukemia (MEL) hybrid cells, the beta-globin gene from the MEL parent is transcriptionally inactivated and is later replicating than in the parental MEL cell line. In human fibroblast (GM3552) x MEL hybrid cells, the human beta-globin gene is transcriptionally activated, and all of the sequences within the human beta-globin domain (200 kilobases) we have examined appear to be earlier replicating than those in the parental fibroblast cell line. The chromatin configuration of the activated human beta-globin domain in the hybrids is relatively more sensitive to nucleases than that in the fibroblasts. Furthermore, major nuclease-hypersensitive sites that were absent in the chromatin flanking the distal 5' region of the human beta-globin gene cluster in the parental fibroblast cell line are present in the transcriptionally activated domain in the hybrid cell line. These results suggest that timing of replication of globin genes has been altered in these hybrid cells and thus is not fixed during the process of differentiation.

Animals↗
PubMed1988

Timing of chromosomal replication in Escherichia coli.

We have previously shown that certain mutations in the dnaA and recA genes of Escherichia coli perturb initiation of chromosomal replication so that all origins present are not initiated simultaneously. In this work, several genes whose protein products are involved in initiation of replication have been investigated for their effects on the synchrony of initiation. Some of the mutants (dnaC2, rpoC907, dam3) were found to have the asynchrony phenotype. Also, dnaA(Ts) mutations were shown to be dominant over dnaA+ in terms of initiation synchrony. The mechanism leading to the asynchronous phenotype is discussed.

Bacterial Proteins↗
PubMed1986

Zinc oxide to induce molt in layers.

The effects of the addition of Zn as ZnO to diets to induce molt were evaluated against a fasted control. Experiment 1 involved 315 Leghorn hens, 15 months old, randomly distributed among five treatments, each replicated seven times with 9 hens per replicate. Hens fasted for 10 days were compared with hens fed diets to which ZnO was added at 10,000, 5,000, or 2,500 ppm for 7, 14, or 21 days. No significant differences were observed among treatments for days to return to 50% production, hen-day and hen-housed production, egg weight, grams egg per hen-day, grams of feed per gram egg, mortality, or Haugh units during the 22-week experimental period. Experiment 2 involved 420 Leghorn hens, 18 months old, randomly distributed among five treatments, each replicated seven times with 12 hens per replicate. Treatments involved fasting for 10 days or feeding diets with 10,000, 5,000, or 2,500 ppm ZnO fed for 7, 14, or 21 days. Hens fasted and hens fed diets with 10,000 ppm ZnO at the start of the experiment ceased production in significantly less time (4.6 to 6 days) than hens fed 5,000 ppm ZnO (14.3 to 14.9 days); however, days to return to 50% production from the start of the experiment did not differ among treatments. Feed consumption and feed cost per hen day during molt were lowest (P less than .05) in the fasted hens.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1998

Parental alleles of an imprinted mouse transgene replicate synchronously.

Molecular features of imprinted genes include differences in expression, methylation, and the timing of DNA replication between parental alleles. Whereas methylation differences always seem to be associated with differences in expression, differences in the timing of replication between parental homologs are not always seen at imprinted loci. These observations raise the possibility that differences in replication timing may not be an essential feature underlying genomic imprinting. In this study, we examined the timing of replication of the two alleles of the imprinted RSVIgmyc transgene in individual embryonic cells using fluorescence in situ hybridization (FISH). The cis-acting signals for RSVIgmyc imprinting are within RSVIgmyc itself. Thus, allele-specific differences in replication, if they indeed govern RSVIgmyc imprinting, should be found in RSVIgmyc sequences. We found that the parental alleles of RSVIgmyc, which exhibit differences in methylation, replicated at the same time. Synchronous replication was also seen in embryonic cells containing a modified version of RSVIgmyc that exhibited parental allele differences in both methylation and expression. These findings indicate that maintenance of expression and methylation differences between alleles does not require a difference in replication timing. The differences in replication timing of endogenous imprinted alleles detected by FISH might therefore reflect structural differences between the two alleles that could be a consequence of imprinting or, alternatively, could be unrelated to imprinting.

Alleles↗
PubMed1988

Heterochromatin associated with active versus inactive centromeres of mouse replicates at different times.

A subline of mouse L-cells carries a dicentric chromosome in which one centromere always separates prematurely. This centromere is not involved in the dynamics of chromosome migration and is considered inactive. By use of anti-BRdU antibody binding to BRdU-treated chromosomes it is shown that the pericentric constitutive heterochromatin associated with the prematurely separating centromere replicates earlier than its counterpart associated with the active centromere and even before several euchromatic regions in the genome. These results point to a possible mechanism by which dicentric chromosomes segregate equationally.

Animals↗
PubMed1996

A meiosis-specific protein kinase, Ime2, is required for the correct timing of DNA replication and for spore formation in yeast meiosis.

In this report we study the regulation of premeiotic DNA synthesis in Saccharomyces cerevisiae. DNA replication was monitored by fluorescence-activated cell sorting analysis and by analyzing the pattern of expression of the DNA polymerase alpha-primase complex. Wild-type cells and cells lacking one of the two principal regulators of meiosis, Ime1 and Ime2, were compared. We show that premeiotic DNA synthesis does not occur in ime1 delta diploids, but does occur in ime2 delta diploids with an 8-9 h delay. At late meiotic times, ime2 delta diploids exhibit an additional round of DNA synthesis. Furthermore, we show that in wild-type cells the B-subunit of DNA polymerase alpha is phosphorylated during premeiotic DNA synthesis, a phenomenon that has previously been reported for the mitotic cell cycle. Moreover, the catalytic subunit and the B-subunit of DNA polymerase alpha are specifically degraded during spore formation. Phosphorylation of the B-subunit does not occur in ime1 delta diploids, but does occur in ime2 delta diploids with an 8-9 h delay. In addition, we show that Ime2 is not absolutely required for commitment to meiotic recombination, spindle formation and nuclear division, although it is required for spore formation.

Cell Cycle Proteins↗
PubMed2005

Bacillus subtilis YabA is involved in determining the timing and synchrony of replication initiation.

It is shown here by flow cytometry that Bacillus subtilis YabA negatively regulates the timing of replication initiation. When the level of YabA was reduced, replication began at a decreased cell mass and when the level was increased, initiation was delayed. Synchrony of replication initiation was also disrupted at low levels of YabA. Yfp-YabA localized as foci in cells. Since YabA was reported to interact with DnaN (beta subunit of DNA polymerase III), co-localization of Yfp-YabA with the polymerase was examined using a Cfp fusion with DnaX (tau subunit of DNA polymerase III). It is reported that YabA appears to localize at the replication forks only at a late stage of DNA replication.

Bacillus subtilis↗
PubMed2003

Replicate real-time PCR testing of DNA in maternal plasma increases the sensitivity of non-invasive fetal sex determination.

BACKGROUND: We determined fetal sex in pregnancies referred for invasive prenatal diagnosis procedures by analysis of DNA in maternal plasma. METHODS: Twelve pregnancies at risk of X-linked haemophilia and 32 pregnancies at risk of chromosomal aneuploidies at a gestational age ranging from 10 to 18 weeks recruited before chorionic villus sampling or amniocentesis were involved in the study. Male fetal DNA in maternal plasma was detected by using real-time polymerase chain reaction with the SRY gene as a marker. RESULTS: The specificity of the system reached 100% (no Y signal was detected in 17 women pregnant with a female fetus) and the sensitivity reached 100% (SRY amplification in 27 examined samples). CONCLUSIONS: Amplification of free fetal DNA in maternal plasma is a valid and rapid technique for predicting fetal sex in first- and second-trimester pregnancies and could allow the restriction of invasive sampling procedures to male fetuses at risk of X-linked disorders.

Aneuploidy↗
PubMed2003

Effects of dietary protein and energy concentrations on performance and carcase characteristics of chukar partridge (Alectoris chukar) raised in captivity.

1. This study was conducted to determine the effects of starter and grower diets with differing crude protein (CP) and metabolisable energy (ME) concentrations on the body weight (BW), live weight gain (LWG), feed consumption (FC), feed conversion ratio (FCR), and carcase, breast+back, rump, wing, neck and abdominal fat weights of chukar partridge raised in captivity. 2. Chukar partridges were fed on starter diets containing 4 concentrations of CP (160, 200, 240, 280 g/kg) and 4 concentrations of ME (10.9, 11.7, 12.6, 13.4 MJ/kg) from hatch to 8 weeks of age; they were fed on grower diets containing 4 concentrations of CP (150, 175, 200, 225 g/kg) and 4 concentrations of ME (11.9, 12.6, 13.2, 13.8 MJ/kg) from 9 to 16 weeks of age. All diets contained at least 5.5 g/kg methionine, 15 g/kg lysine and 10 g/kg methionine+cystine. Sixteen starter and 16 grower diets were arranged in a 4 x 4 factorial design with 4 levels of CP and 4 levels of ME. Each treatment was replicated three times with each replicate consisting of 5 males and 5 females. 3. Partridges fed on a starter diet containing 160 g CP/kg were significantly lighter at 8 weeks of age than those in groups given diets containing a higher CP. However, at 16 weeks of age, the differences in BW among treatments had disappeared. Throughout, there were no significant effects of ME concentration on BW and LWG. 4. The daily mean FC for the 0 to 8 week and 0 to 16 week periods was not affected by dietary CP concentration. For the 9 to 16 week period, the partridges fed on a grower diet containing 225 g CP/kg consumed more feed than those given a diet containing 175 g CP/kg. 5. The highest FCR for the 0 to 8 week period was in partridges fed on a starter diet containing 160 g CP/kg. For the 9 to 16 week period, the lowest FCR was in partridges fed on a grower diet containing 150 g CP/kg. For the 0 to 16 week period, there was not a significant effect of dietary CP concentration on FCR. The daily mean FC and the FCR for the 0 to 8, 9 to 16 and 0 to 16 week periods decreased when the ME concentration of the starter and grower diets increased. 6. The carcase, rump and breast+back weights of the male partridges increased when the ME content of the diets increased. Weights of all carcase components of the male partridges were significantly greater than those of the carcase components of the females. 7. There were no significant interactions between CP and ME concentrations on BW, LWG, FC, FCR and carcase characteristics. 8. We conclude that the starter diet for chukar partridges raised for meat production should contain at least 200 g CP/kg, 11.7 MJ ME/kg, and the grower diet should contain 150 g CP/kg, 12.6 MJ ME/kg.

Age Factors↗
PubMed1994

Polydnavirus of the parasitic wasp Chelonus inanitus (Braconidae): characterization, genome organization and time point of replication.

Ultrastructural analysis of the polydnavirus of the braconid wasp Chelonus inanitus revealed that virions consist of one cylindrical nucleocapsid enveloped by a single unit membrane. Nucleocapsids have a constant diameter of 33.7 +/- 1.4 nm and a variable length of between 8 and 46 nm. Spreading of viral DNA showed that the genome consists of circular dsDNA molecules of variable sizes and measurement of the contour lengths indicated sizes of between 7 and 31 kbp. When virions were exposed to osmotic shock conditions to release the DNA, only one circular molecule was released per particle suggesting that the various DNA molecules are singly encapsidated in this bracovirus. The viral genome was seen to consist of at least 10 different segments and the aggregate genome size is in the order of 200 kbp. By partial digestion of viral DNA with HindIII or EcoRI in the presence of ethidium bromide and subsequent ligation with HindIII-cut pSP65 or EcoRI-cut pSP64 and transfection into Escherichia coli, libraries of 103 HindIII and 23 EcoRI clones were obtained. Southern blots revealed that complete and unrearranged segments were cloned with this approach, and restriction maps for five segments were obtained. Part of a 16.8 kbp segment was sequenced, found to be AT-rich (73%) and to contain six copies of a 17 bp repeated sequence. The development of the female reproductive tract in the course of pupal-adult development of the wasp was investigated and seen to be strictly correlated with the pigmentation pattern. By the use of a semiquantitative PCR, replication of viral DNA was observed to initiate at a specific stage of pupal-adult development.

Animals↗
PubMed1984

Performance, ruminal and serum characteristics of steers fed lasalocid on pasture.

Seventy-two growing steers were used in a 126-d study to determine the influence of varying levels of lasalocid on performance, ruminal and serum characteristics of animals grazing pasture. Treatments consisted of: 1) control; 2) 200 mg lasalocid/d and 3) 300 mg lasalocid/d. Each treatment was replicated three times and each replicate of eight steers was maintained on 3.0 ha of pasture. Pastures consisted of a mixture of tall fescue, orchard grass and ladino clover. In addition to pasture, each replicate of steers was group fed ground corn at a rate of .91 kg X head-1 X d-1 with the lasalocid incorporated into the grain. Average daily gains were .50, .60 and .57 kg, respectively, for steers on the control, and for the 200 and 300 mg lasalocid treatments, which differed (P less than .05) from controls. Ruminal acetate (mol/100 mol) was lower (P less than .05) in steers fed lasalocid at 28 d, but similar for all treatments at 56 and 112 d. Molar proportion of propionate was higher (P less than .05) and butyrate and valerate were lower (P less than .10) in rumen fluid of steers receiving 200 or 300 mg/d of lasalocid. Plasma glucose concentrations were similar for controls and steers receiving 200 mg lasalocid/d, but higher (P less than .05) in steers fed 300 mg lasalocid/d. Serum Mg concentrations were lower (P less than .01) in steers receiving lasalocid. Potassium concentrations in serum were slightly lower (P less than .01) in animals fed lasalocid at 112 d, but not at 28 or 56 d.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗