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1.
Sukhija K Pyne M Ali S Orr V Abedi D Moo-Young M Chou CP 《Molecular biotechnology》2012,51(2):109-118
Most existing genomic engineering protocols for manipulation of Escherichia coli are primarily focused on chromosomal gene knockout. In this study, a simple but systematic chromosomal gene knock-in method
was proposed based on a previously developed protocol using bacteriophage λ (λ Red) and flippase–flippase recognition targets
(FLP–FRT) recombinations. For demonstration purposes, DNA operons containing heterologous genes (i.e., pac encoding E. coli penicillin acylase and palB2 encoding Pseudozyma antarctica lipase B mutant) engineered with regulatory elements, such as strong/inducible promoters (i.e., P
trc
and P
araB
), operators, and ribosomal binding sites, were integrated into the E. coli genome at designated locations (i.e., lacZYA, dbpA, and lacI-mhpR loci) either as a gene replacement or gene insertion using various antibiotic selection markers (i.e., kanamycin and chloramphenicol)
under various genetic backgrounds (i.e., HB101 and DH5α). The expression of the inserted foreign genes was subjected to regulation
using appropriate inducers [isopropyl β-d-1-thiogalactopyranoside (IPTG) and arabinose] at tunable concentrations. The developed approach not only enables more extensive
genomic engineering of E. coli, but also paves an effective way to “tailor” plasmid-free E. coli strains with desired genotypes suitable for various biotechnological applications, such as biomanufacturing and metabolic
engineering. 相似文献
2.
In the developing peanut (Arachis hypogaea L.) kernels, the period between 15 and 35 days after podding (DAP) was identified as the active period of oil-filling. The
period of active oil-filling was associated with a decrease in the starch, soluble sugars and proteins so as to make available
the energy and carbon skeleton for the synthesis of oil. The oil content in the mature kernels decreased by 11, 12 and 25
per cent with Zn, S and Zn+S deficiency, respectively. In addition, proteins and starch content decreased significantly while
that of soluble sugars increased slightly. The activity of malate dehydrogenase and glucose-6-phosphate dehydrogenase also
decreased due to Zn as well as S deficiency. The deficiency treatments resulted in a decrease in phospholipids, free fatty
acids and triacylglycerols in mature kernels. Further the proportion of 16∶0 and 18∶2 decreased while that of 18∶1 increased
in developing kernels. 相似文献
3.
KB Cullberg T Christiansen SK Paulsen JM Bruun SB Pedersen B Richelsen 《Obesity (Silver Spring, Md.)》2013,21(3):454-460
Background:
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4.
5.
P.J. Niziolek S. Murthy S.N. Ellis K.B. Sukhija T.A. Hornberger C.H. Turner A.G. Robling 《Journal of cellular physiology》2009,221(3):579-585
The osteo‐anabolic effects of intermittent parathyroid hormone (PTH) treatment require insulin‐like growth factor (IGF) signaling through the IGF‐I receptor. A major downstream target of the IGF‐I receptor (via Akt) is the mammalian target of rapamycin (mTOR), a kinase involved in protein synthesis. We investigated whether the bone‐building effects of intermittent PTH require functional mTOR signaling. Mice were treated with daily PTH 1–34 (0, 10, 30, or 90 µg/kg) for 6 weeks in the presence or absence of rapamycin, a selective inhibitor of mTOR. We found that all PTH doses were effective in enhancing bone mass, whether rapamycin was present or not. Rapamycin had little to no effect on the anabolic response at low (10 µg) PTH doses, small effects in a minority of anabolic measures at moderate doses (30 µg), but the anabolic effects of high‐dose PTH (90 µg) were consistently and significantly suppressed by rapamycin (~4–36% reduction). Serum levels of Trap5b, a marker of resorption, were significantly enhanced by rapamycin, but these effects were observed whether PTH was absent or present. Our data suggest that intermittent PTH, particularly at lower doses, is effective in building bone mass in the presence of rapamycin. However, the full anabolic effects of higher doses of PTH are significantly suppressed by rapamycin, suggesting that PTH might normally activate additional pathways (including mTOR) for its enhanced high‐dose anabolic effects. Clinical doses of intermittent PTH could be an effective treatment for maintaining or increasing bone mass among patients taking rapamycin analogs for unrelated health issues. J. Cell. Physiol. 221: 579–585, 2009. © 2009 Wiley‐Liss, Inc. 相似文献
6.
7.
KB Gudmundsdóttir S Sigurdarson J Kristinsson T Eiríksson T Jóhannesson 《Acta veterinaria Scandinavica》2006,48(1):16-5
This study was undertaken in order to examine whether any connection existed between the amounts of iron in forage and the
sporadic occurrence of scrapie observed in certain parts of Iceland. As iron and manganese are considered antagonistic in
plants, calculation of the Fe/Mn ratios was also included by using results from Mn determination earlier performed in the
same samples. Forage samples (n = 170) from the summer harvests of 2001–2003, were collected from 47 farms for iron and manganese
analysis. The farms were divided into four categories: 1. Scrapie-free farms in scrapie-free areas (n = 9); 2. Scrapie-free farms in scrapie-afflicted areas (n = 17); 3. Scrapie-prone farms (earlier scrapie-afflicted, restocked farms) (n = 12); 4. Scrapie-afflicted farms (n = 9). Farms in categories 1 and 2 are collectively referred to as scrapie-free farms. The mean iron concentration in forage samples from scrapie-afflicted farms was significantly higher than in forage samples
from farms in the other scrapie categories (P = 0.001). The mean Fe/Mn ratio in forage from scrapie-afflicted farms was significantly
higher than in forage from scrapie-free and scrapie-prone farms (P < 0.001). The results indicated relative dominance of iron
over manganese in forage from scrapie-afflicted farms as compared to farms in the other categories. Thus thorough knowledge
of iron, along with manganese, in soil and vegetation on sheep farms could be a pivot in studies on sporadic scrapie. 相似文献
8.
Seyung Chung Sami Dwabe Yayha Elshimali Hemlata Sukhija Clement Aroh Jaydutt V. Vadgama 《PloS one》2015,10(8)
Colorectal cancer (CRC) is one of the three leading causes for cancer mortality. CRC kills over 600,000 people annually worldwide. The most common cause of death from CRC is the metastasis to distant organs. However, biomarkers for CRC metastasis remain ill-defined. We compared primary and metastatic CRC cell lines for their angiogenesis-protein profiles and intracellular signaling profiles to identify novel biomarkers for CRC metastasis. To this end, we used primary and metastatic CRC cell lines as a model system and normal human colon cell line as a control. The angiogenesis profiles two isogenic CRC cell lines, SW480 and SW620, and HT-29 and T84 revealed that VEGF was upregulated in both SW620 and T84 whereas coagulation factor III, IGFBP-3, DPP IV, PDGF AA/AB, endothelin I and CXCL16 were downregulated specifically in metastatic cell lines. Furthermore, we found that TIMP-1, amphiregulin, endostatin, angiogenin were upregulated in SW620 whereas downregulated in T84. Angiogenin was downregulated in T84 and GM-CSF was also downregulated in SW620. To induce CRC cell metastasis, we treated cells with pro-inflammatory cytokine IL-6. Upon IL-6 treatment, epithelial-mesenchymal transition was induced in CRC cells. When DLD-1 and HT-29 cells were treated with IL-6; Akt, STAT3, AMPKα and Bad phosphorylation levels were increased. Interestingly, SW620 showed the same signal activation pattern with IL-6 treatment of HT-29 and DLD-1. Our data suggest that Akt, STAT3, AMPKα and Bad activation can be biomarkers for metastatic colorectal cancer. IL-6 treatment specifically reduced phosphorylation levels of EGFR, HER2 receptor, Insulin R and IGF-1R in receptor tyrosine kinase array study with HT-29. Taken together, we have identified novel biomarkers for metastatic CRC through the angiogenesis-antibody array and intracellular signaling array studies. Present study suggests that those novel biomarkers can be used as CRC prognosis biomarkers, and as potential targets for the metastatic CRC therapy. 相似文献
9.
Scanning electron microscopy and gramicidin patch clamp recordings of microvillous receptor neurons dissociated from the rat vomeronasal organ 总被引:2,自引:1,他引:1
Vomeronasal organs from female rats were dissociated and isolated
microvillous receptor neurons were studied. The isolated receptor neurons
kept the typical bipolar shape which they have in situ as observed by
scanning electron microscopy. We applied the perforated patch-clamp
technique using the cation-selective ionophore gramicidin on freshly
isolated and well differentiated receptor neurons. The mean resting
potential was -58+/-14 mV (n=39). The contribution of the sodium pump
current to the resting potential was demonstrated by lowering the K+
concentration in the bath or by application of 100 microM dihydro-ouabain.
The input resistance was in the range of 1-6 GOmega and depolarizing
current pulses of a few pA were sufficient to trigger overshooting action
potentials. In voltage clamp conditions a fast transient sodium inward
current and a sustained outward potassium current were activated by
membrane depolarization. These observations indicate that freshly isolated
vomeronasal receptor neurons of rats can be recorded, using gramicidin,
with little modification of the intracellular content. Their
electrophysiological properties are very similar to those observed in situ.
Four out of eight female vomeronasal receptor cells were depolarized by
diluted rat male urine.
相似文献
10.