| 1. |
- Beal, Jacob, et al.
(författare)
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Robust estimation of bacterial cell count from optical density
- 2020
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Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
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| 5. |
- Sodergren, Erica, et al.
(författare)
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The genome of the sea urchin Strongylocentrotus purpuratus.
- 2006
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Ingår i: Science. - : American Association for the Advancement of Science (AAAS). - 1095-9203 .- 0036-8075. ; 314:5801, s. 941-52
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Tidskriftsartikel (refereegranskat)abstract
- We report the sequence and analysis of the 814-megabase genome of the sea urchin Strongylocentrotus purpuratus, a model for developmental and systems biology. The sequencing strategy combined whole-genome shotgun and bacterial artificial chromosome (BAC) sequences. This use of BAC clones, aided by a pooling strategy, overcame difficulties associated with high heterozygosity of the genome. The genome encodes about 23,300 genes, including many previously thought to be vertebrate innovations or known only outside the deuterostomes. This echinoderm genome provides an evolutionary outgroup for the chordates and yields insights into the evolution of deuterostomes.
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| 6. |
- James, Katherine L., et al.
(författare)
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Low-Bias RNA Sequencing of the HIV-2 Genome from Blood Plasma
- 2019
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Ingår i: Journal of Virology. - 1098-5514. ; 93:1
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Tidskriftsartikel (refereegranskat)abstract
- Accurate determination of the genetic diversity present in the HIV quasispecies is critical for the development of a preventative vaccine: in particular, little is known about viral genetic diversity for the second type of HIV, HIV-2. A better understanding of HIV-2 biology is relevant to the HIV vaccine field because a substantial proportion of infected people experience long-term viral control, and prior HIV-2 infection has been associated with slower HIV-1 disease progression in coinfected subjects. The majority of traditional and next-generation sequencing methods have relied on target amplification prior to sequencing, introducing biases that may obscure the true signals of diversity in the viral population. Additionally, target enrichment through PCR requires a priori sequence knowledge, which is lacking for HIV-2. Therefore, a target enrichment free method of library preparation would be valuable for the field. We applied an RNA shotgun sequencing (RNA-Seq) method without PCR amplification to cultured viral stocks and patient plasma samples from HIV-2-infected individuals. Libraries generated from total plasma RNA were analyzed with a two-step pipeline: (i) de novo genome assembly, followed by (ii) read remapping. By this approach, whole-genome sequences were generated with a 28× to 67× mean depth of coverage. Assembled reads showed a low level of GC bias, and comparison of the genome diversities at the intrahost level showed low diversity in the accessory gene vpx in all patients. Our study demonstrates that RNA-Seq is a feasible full-genome de novo sequencing method for blood plasma samples collected from HIV-2-infected individuals.IMPORTANCE An accurate picture of viral genetic diversity is critical for the development of a globally effective HIV vaccine. However, sequencing strategies are often complicated by target enrichment prior to sequencing, introducing biases that can distort variant frequencies, which are not easily corrected for in downstream analyses. Additionally, detailed a priori sequence knowledge is needed to inform robust primer design when employing PCR amplification, a factor that is often lacking when working with tropical diseases localized in developing countries. Previous work has demonstrated that direct RNA shotgun sequencing (RNA-Seq) can be used to circumvent these issues for hepatitis C virus (HCV) and norovirus. We applied RNA-Seq to total RNA extracted from HIV-2 blood plasma samples, demonstrating the applicability of this technique to HIV-2 and allowing us to generate a dynamic picture of genetic diversity over the whole genome of HIV-2 in the context of low-bias sequencing.
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| 7. |
- Obregon-Whittle, Maria V, et al.
(författare)
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Enamel matrix derivative stimulates expression and secretion of resistin in mesenchymal cells.
- 2011
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Ingår i: European journal of oral sciences. - : Wiley. - 1600-0722 .- 0909-8836. ; 119 Suppl 1, s. 366-72
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Tidskriftsartikel (refereegranskat)abstract
- In this study we wanted to identify the effect of enamel matrix derivative (EMD) on adipocytokines, so-called adipokines. Primary human cells of mesenchymal origin (osteoblasts, periodontal ligament cells, mesenchymal stem cells, and pulp cells) and hematopoietic origin (monocytes) were incubated with EMD. The levels of adipokines in cell culture medium were quantified using the Lincoplex human adipocyte panel (Luminex) and by real-time PCR of mRNA isolated from cell lysates. Rats were injected with 2 mg of EMD or saline intramuscularly every third day for 14 d. Blood samples were taken before and after injections, and the level of resistin in rat plasma was measured by ELISA. We found a dramatic increase in the secretion of resistin from mesenchymal stem cells, and verified this result in all the cells of mesenchymal origin tested. However, we observed no significant changes in the amount of resistin secreted from monocytes exposed to EMD compared with the control. Injections of EMD significantly enhanced the circulating levels of resistin in rats, and EMD also significantly enhanced the activity of the resistin promoter in transfected mesenchymal stem cells, indicating a direct effect on resistin expression. Our results indicate that resistin may play a role in mediating the biological effect of EMD in mesenchymal tissues.
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| 8. |
- Whittle, Thomas A., et al.
(författare)
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Tailoring phase transition temperatures in perovskites via A-site vacancy generation
- 2017
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Ingår i: Dalton Transactions. - : Royal Society of Chemistry (RSC). - 1477-9226 .- 1477-9234. ; 46:22, s. 7253-7260
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Tidskriftsartikel (refereegranskat)abstract
- The structures across the Sr0.8Ti0.6-xZrxNb0.4O3, 0 <= x <= 0.6, defect perovskite series were investigated using complementary synchrotron X-ray and neutron powder diffraction data. The locations of second order compositional and temperature dependent phase transitions between the high symmetry cubic Pm (3) over barm phase and the lower symmetry tetragonal I4/mcm phase were determined. Deviation of the oxygen x coordinate from the high symmetry value and the B-O-B bond angle from 180 degrees as well as the tetragonal strain squared were each found to be suitable order parameters to monitor the transitions. Tolerance factor calculations confirmed that these A-site deficient perovskites retain a higher symmetry to a lower value than their fully occupied counterparts. Therefore, adjusting vacancy concentrations can be employed as a general strategy to design compounds with specifically tailored phase transition temperatures.
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| 9. |
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- 2017
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Ingår i: Physical Review D. - 2470-0010 .- 2470-0029. ; 96:2
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Tidskriftsartikel (refereegranskat)
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