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2.
The site specificity of bacteriophage T4-induced type II DNA topoisomerase action on double-stranded DNA has been explored by studying the sites where DNA cleavages are induced by the enzyme. Oxolinic acid addition increases the frequency at which phi X174 duplex DNA is cut by the enzyme by about 100-fold, to the point where nearly every topoisomerase molecule causes a double-stranded DNA cleavage event. The effect of oxolinic acid on the enzyme is very similar to its effect on another type II DNA topoisomerase, the Escherichia coli DNA gyrase. A filter-binding method was developed that allows efficient purification of topoisomerase-cleaved DNA fragments by selecting for the covalent attachment of this DNA to the enzyme. Using this method, T4 topoisomerase recognition of mutant cytosine-containing T4 DNA was found to be relatively nonspecific, whereas quite specific recognition sites were observed on native T4 DNA, which contains glucosylated hydroxymethylcytosine residues. The increased specificity of native T4 DNA recognition seems to be due entirely to the glucose modification. In contrast, E. coli DNA gyrase shows a high level of specificity for both the mutant cytosine-containing DNA and native T4 DNA, recognizing about five strong cleavage sites on both substrates. An unexpected feature of DNA recognition by the T4 topoisomerase is that the addition of the cofactor ATP strongly stimulates the topoisomerase-induced cleavage of native T4 DNA, but has only a slight effect on cleavage of cytosine-containing T4 DNA.  相似文献   
3.
Catecholamines in plasma and urine at high altitude   总被引:4,自引:0,他引:4  
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4.
The rolling circle DNA replication structures generated by the in vitro phage T4 replication system were analyzed using two-dimensional agarose gels. Replication structures were generated in the presence or absence of T4 primase (gp61), permitting the analysis of replication forks with either duplex or single-stranded tails. A characteristic arc shape was visualized when forks with single-stranded tails were cleaved by a restriction enzyme with the help of an oligonucleotide that anneals to restriction sites in the single-stranded tail. After calibrating the gel system with this well-studied rolling circle replication reaction, we then analyzed the in vivo replication directed by a T4 replication origin cloned within a plasmid. DNA samples were generated from infections with either wild-type or primase-deletion mutant phage. The only replicative arc that could be detected in the wild-type sample corresponded to duplex Y forms, consistent with very efficient lagging strand synthesis. Surprisingly, we obtained evidence for both duplex and single-stranded DNA tails in the samples from the primase-deficient infection. We conclude that a relatively inefficient mechanism primes lagging strand DNA synthesis in vivo when gp61 is absent.  相似文献   
5.
Six hundred and two mammary tumors were examined clinically, by mammography and cytology, with a histologic checkup following surgical biopsy. There were 247 cases of malignoma and 355 benign cases. The limited reliability of the individual methods is demonstrated, and it is shown that their combined use can improve the diagnosis. More malignomas are detected, and preoperative diagnosis is made more safely. If all three methods yield identical results, as was the case in 50.2% of the malignant and 32.7% of the benign lesions, the probability of diagnostic error is less than one per cent. With a malignoma thus established, surgical treatment may follow immediately, or irradiation can be started. In benign cases a surgical biopsy may be foregone and further developments may be awaited with due provision for regular control. If the three methods yield conflicting or doubtful results, elucidation by surgical biopsy and histology is indicated.  相似文献   
6.
Recent advances in the area of bacterial DNA damage responses are reviewed here. The SOS pathway is still the major paradigm of bacterial DNA damage response, and recent studies have clarified the mechanisms of SOS induction and key physiological roles of SOS including a very major role in genetic exchange and variation. When considering diverse bacteria, it is clear that SOS is not a uniform pathway with one purpose, but rather a platform that has evolved for differing functions in different bacteria. Relating in part to the SOS response, the field has uncovered multiple apparent cell-cycle checkpoints that assist cell survival after DNA damage and remarkable pathways that induce programmed cell death in bacteria. Bacterial DNA damage responses are also much broader than SOS, and several important examples of LexA-independent regulation will be reviewed. Finally, some recent advances that relate to the replication and repair of damaged DNA will be summarized.Since the publication of DNA Repair and Mutagenesis in 2006 (Friedberg et al. 2006), our understanding of bacterial DNA damage responses has progressed significantly. Some studies have refined known pathways and filled in important details, whereas other studies have uncovered surprising new pathways such as bacterial programmed cell death and a form of replicative repair that reconstitutes severely shattered genomes. This review will focus on these recent advances, with only limited discussion and citation to work that precedes the 2006 tome.  相似文献   
7.
Integration of Plasmids into the Bacteriophage T4 Genome   总被引:2,自引:0,他引:2       下载免费PDF全文
HWE. Kreuzer  K. N. Kreuzer 《Genetics》1994,138(4):983-992
We have analyzed the integration of plasmids into the bacteriophage T4 genome via homologous recombination. As judged by genetic selection for a plasmid-borne marker, a mutation in phage gene uvsX or uvsY essentially blocked the integration of a plasmid with homology to the T4 genome but no phage replication origin (non-origin plasmid). The strict requirement for these two proteins suggests that plasmid integration can proceed via a strand-invasion reaction similar to that catalyzed in vitro by the T4-encoded strand-exchange protein (UvsX) in concert with UvsY and gp32. In contrast to the results with the non-origin plasmid, a mutation in uvsX or uvsY reduced the integration of a T4 replication origin-containing plasmid by only 3-10-fold. These results suggest that the origin-containing plasmid integrates by both the UvsXY-dependent pathway used by the non-origin plasmid and by a UvsXY-independent pathway. The origin-containing plasmid integrated into the phage genome during a uvsX-or uvsY-mutant infection of a recA-mutant host, and therefore origin-dependent integration can occur in the absence of both phage- and host-encoded strand-exchange proteins (UvsX and RecA, respectively).  相似文献   
8.
The aim of the present experiment was to compare silage prepared from maize having a brown midrib (BMR) mutation with control (CTR) maize to identify their effects on enteric methane emission, digesta mean retention time (MRT), ruminal fermentation and digestibility. In addition, the utility of archaeol present in faecal samples was validated as a proxy for methane production. Seven German Holstein heifers were fed total mixed rations with a maize-silage proportion (either BMR or CTR) of 920 g/kg dry matter (DM) in a change-over design. Heifers were fed boluses with markers to measure MRT; faeces were collected for 7 days and rumen fluid was collected on the penultimate day. Methane emission was measured in respiration chambers on one day. Data were analysed by t-test and regression analysis. DM intake did not differ between the two diets. The apparent digestibility of DM and most nutrients was unaffected by diet type, but apparent digestibility of neutral and acid detergent-fibre was higher in those heifers fed BMR than in those fed CTR. Comparisons between diets revealed no difference in particle or solute MRT in the gastro-intestinal tract and the reticulorumen. Concentrations of short-chain fatty acid and ammonia in rumen fluid and its pH were not affected by silage type. Independent of the mode of expression [l/d, l/kg DM intake, l/kg digested organic matter], methane emissions were not affected by maize-silage type, but with BMR, there was a trend towards lower methane production per unit of digested neutral detergent fibre than there was with CTR silage. Results of the present study show that feeding heifers BMR silage does not increase methane emissions despite a higher fibre digestibility as compared to CTR silage. Therefore, it is assumed that improvements in animal productivity achieved by feeding BMR silage, as some studies have reported, can be obtained without extra environmental cost per unit of milk or meat. Neither faecal archaeol content [µg/g] nor daily amount excreted [mg/d] is suitable to predict methane production in absolute terms [l per day]. However, faecal archaeol content has a certain potential for predicting the methane yield [l per kg DM intake] of individual animals.  相似文献   
9.
The effects of tanniferous sainfoin on digestion and metabolism have been investigated in 12 lambs in an incomplete cross-over design (n = 6). Effects of condensed tannins (CT) were evaluated by comparing dehydrated and ensiled sainfoin treated with and without polyethylene glycol (PEG). Dehydrated and ensiled grass-clover mixtures served as controls. The lambs were fed the treatment diets, including a mineral supplement, for 21 d. During the last 7 d excreta, rumen fluid and blood were sampled. The CT of sainfoin decreased rumen fluid ammonia concentration (p < 0.001) and increased the plasma concentration mainly of essential amino acids (p < 0.001). Body retention of phosphorus, calcium and magnesium was lower with sainfoin compared to PEG-treated sainfoin (p < 0.05). Sainfoin without PEG resulted in lower digestibilities of organic matter and neutral detergent fibre than sainfoin with PEG and the grass-clover mixture (p < 0.001). Ensiling of sainfoin led to the lowest N-retention. In conclusion, the reduction in ruminal ammonia and urine-N losses by sainfoin CT did not improve N-retention.  相似文献   
10.
Buckwheat is of high value in crop rotations and overall agricultural ecology because of strong rooting and intensive flowering properties, but it is rarely cultivated and information on its nutritional value to ruminants is scarce. The contents of net energy for lactation (NEL), as estimated with the Hohenheim Gas Test (n = 3), were 4.3, 4.9 and 7.5 MJ NEL/kg dry matter (DM) for fresh and ensiled whole buckwheat plants and buckwheat grain, respectively. In two experiments with the Rumen Simulation Technique (Rusitec), ruminal fermentation characteristics of buckwheat forages and buckwheat grain (n = 4/diet) were evaluated. In the first experiment, 0, 300 or 600 g/kg of a pure hay diet were replaced by either fresh or ensiled buckwheat to create five diets. Neither form of buckwheat forages had effects on in vitro ruminal degradability and short chain fatty acid (SCFA) concentrations and composition. The use of fresh buckwheat reduced ruminal ammonia concentrations and enhanced estimated microbial N growth efficiency. These differences did not occur with silage, indicating a change in nutritional value by ensiling. Fresh buckwheat reduced the number of bacteria in the incubated fluid, while ensiled buckwheat reduced that of holotrich protozoa. Methane formation was not influenced by the buckwheat forages. In the second experiment, wheat meal (400 g/kg dietary DM), was replaced stepwise (0.5 and 1.0) by buckwheat grain meal. This did not cause differences in parameters of nutrient degradability, relative N efficiency and total amount and composition of SCFA. Holotrich protozoa counts increased, but total gas formation decreased with increasing dietary level of buckwheat grain. In a final experiment, cows yielding about 40 kg milk/day were fed mixed silage-concentrate diets (n = 4). A control diet contained no buckwheat. In a second diet, maize silage was partly substituted by buckwheat silage (98 g/kg dietary DM). In a third group, part of the energy concentrate was substituted by buckwheat grain meal (94 g/kg). There were no effects on feed intake, milk yield and milk composition. Buckwheat proved to be a plant that offers different feeds of a quality sufficient to be considered suitable in ruminant nutrition.  相似文献   
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