The signal produced by fluorescence in situ hybridization (FISH) often is inconsistent among cells and sensitivity is low. Small DNA targets on the chromatin are difficult to detect. We report here an improved nick translation procedure for Texas red and Alexa Fluor 488 direct labeling of FISH probes. Brighter probes can be obtained by adding excess DNA polymerase I. Using such probes, a 30?kb yeast transgene, and the rp1, rp3 and zein multigene clusters were clearly detected. 相似文献
Tuberculosis (TB) is an infectocontagious respiratory disease caused by members
of the Mycobacterium tuberculosis complex. A 7 base pair (bp)
deletion in the locus polyketide synthase
(pks)15/1 is described as polymorphic among members of the
M. tuberculosis complex, enabling the identification of
Euro-American, Indo-Oceanic and Asian lineages. The aim of this study was to
characterise this locus in TB isolates from Mexico. One hundred
twenty clinical isolates were recovered from the states of Veracruz and Estado
de Mexico. We determined the nucleotide sequence of a ± 400 bp fragment of the
locus pks15/1, while genotypic characterisation was
performed by spoligotyping. One hundred and fifty isolates contained the 7 bp
deletion, while five had the wild type locus. Lineages X (22%),
LAM (18%) and T (17%) were the most frequent; only three (2%) of the isolates
were identified as Beijing and two (1%) EAI-Manila. The wild type
pks15/1 locus was observed in all Asian lineage isolates
tested. Our results confirm the utility of locus pks15/1 as a
molecular marker for identifying Asian lineages of the M.
tuberculosis complex. This marker could be of great value in the
epidemiological surveillance of TB, especially in countries like Mexico, where
the prevalence of such lineages is unknown. 相似文献
This study evaluated the effect of different doses of the antioxidant α‐lipoic acid (LA) administered by intraperitoneal injection on the detoxifying capacity (activity of glutathione‐S‐transferase, GST) and oxidative damage (lipids and proteins) in the pompano, Trachinotus marginatus. The plasma glucose levels showed that there were no differences between the treatments (P > 0.05). In the brain, GST activity was significantly higher (P < 0.05) in fish injected with 40 mg LA kg?1 when compared with the control group. In the muscle, GST activity was not influenced by LA treatment (P > 0.05). In the liver, fish injected with 20 mg LA kg?1 showed higher GST activity than the control group (P < 0.05); however, higher doses (40 and 60 mg LA kg?1) led to a reduction of GST activity in the liver, which was comparable to that observed in the control group (P > 0.05). The two highest LA doses (40 and 60 mg kg?1) had opposite effects, depending on the tissue examined: LA was an antioxidant in the brain, reducing lipid peroxidation (P < 0.05), and a pro‐oxidant in the liver, augmenting oxidative lipid damage (P < 0.05). The latter effect was accompanied by an increase in the free iron concentration in the liver at higher LA doses. These results indicate the need to thoroughly evaluate the antioxidant effects on aquatic organisms, since at some doses and/or in some organs their beneficial effects can be lost. 相似文献
The molecular integrity of the active site of phytases from fungi is critical for maintaining phytase function as efficient catalytic
machines. In this study, the molecular dynamics (MD) of two monomers of phytase B from Aspergillus niger, the disulfide intact
monomer (NAP) and a monomer with broken disulfide bonds (RAP), were simulated to explore the conformational basis of the
loss of catalytic activity when disulfide bonds are broken. The simulations indicated that the overall secondary and tertiary
structures of the two monomers were nearly identical but differed in some crucial secondary–structural elements in the vicinity of
the disulfide bonds and catalytic site. Disulfide bonds stabilize the β-sheet that contains residue Arg66 of the active site and
destabilize the α-helix that contains the catalytic residue Asp319. This stabilization and destabilization lead to changes in the shape
of the active–site pocket. Functionally important hydrogen bonds and atomic fluctuations in the catalytic pocket change during the
RAP simulation. None of the disulfide bonds are in or near the catalytic pocket but are most likely essential for maintaining the
native conformation of the catalytic site.
Abbreviations
PhyB - 2.5 pH acid phophatese from Aspergillus niger,
NAP - disulphide intact monomer of Phytase B,
RAP - disulphide reduced monomer of Phytase B,
Rg - radius of gyration,
RMSD - root mean square deviation,
MD - molecular dynamics. 相似文献
Vascular growth is a prerequisite for adipose tissue (AT) development and expansion. Some AT cytokines and hormones have effects on vascular development, like vascular endothelial growth factor (VEGF‐A), angiopoietin (ANG‐1), ANG‐2 and angiopoietin‐like protein‐4 (ANGPTL‐4).
Methods:
In this study, the independent and combined effects of diet‐induced weight loss and exercise on AT gene expression and proteins levels of those angiogenic factors were investigated. Seventy‐nine obese males and females were randomized to: 1. Exercise‐only (EXO; 12‐weeks exercise without diet‐restriction), 2. Hypocaloric diet (DIO; 8‐weeks very low energy diet (VLED) + 4‐weeks weight maintenance diet) and 3. Hypocaloric diet and exercise (DEX; 8‐weeks VLED + 4‐weeks weight maintenance diet combined with exercise throughout the 12 weeks). Blood samples and fat biopsies were taken before and after the intervention.
Results:
Weight loss was 3.5 kg in the EXO group and 12.3 kg in the DIO and DEX groups. VEGF‐A protein was non‐significantly reduced in the weight loss groups. ANG‐1 protein levels were significantly reduced 22‐25% after all three interventions (P < 0.01). The ANG‐1/ANG‐2 ratio was also decreased in all three groups (P < 0.05) by 27‐38%. ANGPTL‐4 was increased in the EXO group (15%, P < 0.05) and 9% (P < 0.05) in the DIO group. VEGF‐A, ANG‐1, and ANGPTL‐4 were all expressed in human AT, but only ANGPTL‐4 was influenced by the interventions.
Conclusions:
Our data show that serum VEGF‐A, ANG‐1, ANG‐2, and ANGPTL‐4 levels are influenced by weight changes, indicating the involvement of these factors in the obese state. Moreover, it was found that weight loss generally was associated with a reduced angiogenic activity in the circulation. 相似文献
Background: Prolonged mitotic arrest in response to anti-cancer chemotherapeutics, such as DNA-damaging agents, induces apoptosis, mitotic catastrophe, and senescence. Disruptions in mitotic checkpoints contribute resistance to DNA-damaging agents in cancer. MAD2 has been associated with checkpoint failure and chemotherapy response. In this study, a novel splice variant of MAD2, designated MAD2γ, was identified, and its association with the DNA damage response was investigated.
Methods: Endogenous expression of MAD2γ and full-length MAD2 (MAD2α) was measured using RT-PCR in cancer cell lines, normal foreskin fibroblasts, and tumor samples collected from patients with testicular germ cell tumors (TGCTs). A plasmid expressing MAD2γ was transfected into HCT116 cells, and its intracellular localization and checkpoint function were evaluated according to immunofluorescence and mitotic index.
Results: MAD2γ was expressed in several cancer cell lines and non-cancerous fibroblasts. Ectopically expressed MAD2γ localized to the nucleus and reduced the mitotic index, suggesting checkpoint impairment. In patients with TGCTs, the overexpression of endogenous MAD2γ, but not MAD2α, was associated with resistance to cisplatin-based chemotherapy. Likewise, cisplatin induced the overexpression of endogenous MAD2γ, but not MAD2α, in HCT116 cells.
Conclusions: Overexpression of MAD2γ may play a role in checkpoint disruption and is associated with resistance to cisplatin-based chemotherapy in TGCTs. 相似文献
The Cape Gannet Morus capensis is one of several seabird species endemic to the Benguela upwelling ecosystem (BUS) but whose population has recently decreased, leading to an unfavourable IUCN Red List assessment. Application of ‘JARA’ (‘Just Another Red-List Assessment,’ a Bayesian state-space tool used for IUCN Red List assessments) to updated information on the areas occupied by Cape Gannets and the nest densities of breeding birds at their six colonies, suggested that the species should be classified as Vulnerable. However, the rate of decrease of Cape Gannets in their most-recent generation exceeded that of the previous generation, primarily as a result of large decreases at Bird Island, Lambert’s Bay, and Malgas Island, off South Africa’s west coast (the western part of their range). Since the 1960s, there has been an ongoing redistribution of the species from northwest to southeast around southern Africa, and ~70% of the population now occurs on the south coast of South Africa, at Bird Island in Algoa Bay, on the eastern border of the BUS. Recruitment rather than adult survival may be limiting the present population; however, information on the seabird’s demographic parameters and mortality in fisheries is lacking for colonies in the northern part of the BUS. Presently, major threats to Cape Gannet include: substantially decreased availability of their preferred prey in the west; heavy mortalities of eggs, chicks and fledglings at and around colonies, inflicted by Cape Fur Seals Arctocephalus pusillus and other seabirds; substantial disturbance at colonies caused by Cape Fur Seals attacking adult gannets ashore; oiling; and disease. 相似文献
Plant Cell, Tissue and Organ Culture (PCTOC) - Shake-flask in vitro culture of Buddleja cordata cells produces large amounts of biomass and synthetizes verbascoside (VB), linarin and... 相似文献