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BioTechniques

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https://www.readbyqxmd.com/read/29048272/multiplexing-using-isothermal-recombinase-polymerase-amplification-rpa
#1
(no author information available yet)
No abstract text is available yet for this article.
October 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/29048271/microscale-thermophoresis-mst-for-studying-actin-polymerization-kinetics
#2
Andrea Topf, Peter Franz, Georgios Tsiavaliaris
Here, we present a MicroScale Thermophoresis (MST)-based assay for in vitro assessment of actin polymerization. By monitoring the thermophoretic behavior of ATTO488-labeled actin in a temperature gradient over time, we could follow polymerization in real time and resolve its three characteristic phases: nucleation, elongation, and steady-state equilibration. Titration experiments allowed us to evaluate the effects of actin-binding proteins (ABPs) on polymerization, including DNase I-induced inhibition and mDia2FH1FH2 (mDia2)-assisted acceleration of nucleation...
October 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/29048270/genotyping-live-fish-larvae-non-lethal-and-noninvasive-dna-isolation-from-3-5-day-old-hatchlings
#3
G Janelle Espinoza, J Michael Poland, Jaime R Alvarado Bremer
Genotyping fish larvae is a valuable technique for numerous fields of study. While methods for collecting DNA from early stage larvae have been published, a non-lethal, non-invasive genotyping protocol for hatchlings that is amenable to high-throughput approaches is desirable. Here, we describe a method to individually genotype live, free-swimming, early fish larvae by characterizing their environmental DNA (eDNA). We demonstrate the utility of the method by assigning parentage to a sample (n = 50) of 3-5-day-old sheepshead minnow (Cyprinodon variegatus) larvae hatchlings, with very high rates of genotyping success (98%) and survival (92%) using mitochondrial and microsatellite DNA data...
October 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/29048269/an-improved-collagen-zymography-approach-for-evaluating-the-collagenases-mmp-1-mmp-8-and-mmp-13
#4
Seniz Inanc, Didem Keles, Gulgun Oktay
Collagen zymography is an SDS-PAGE-based method for detecting both the proenzyme and active forms of collagenases. Although collagen zymography is used for assessment of the matrix metalloproteinases MMP-1 and MMP-13, it can be difficult to detect these collagenases due to technical issues. Moreover, it remains unclear whether the collagenase activity of MMP-8 can be detected by this method. Here, we present an improved collagen zymography method that allows quantification of the activities of MMP-1, MMP-8, and MMP-13...
October 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/29048268/density-separation-of-quiescent-yeast-using-iodixanol
#5
Ishtiaque Quasem, Christopher J Luby, Charles R Mace, Stephen M Fuchs
As yeast are starved of nutrients, they enter G0, a quiescent state. Quiescent yeast (Q) cells retain viability for extended periods of time and resume growth following supplementation of missing nutrients. As such, Q cells have become a valuable model for studying longevity and self-renewal of chronologically aged cells. Traditional isolation of Q cells involves a relatively long centrifugation time through a continuous density gradient. Here, we describe a rapid and cost-effective Q-cell isolation technique that uses a single-density, one-step gradient prepared from media containing iodixanol...
October 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/29048267/image-based-cell-size-estimation-for-baculovirus-quantification
#6
T├Ánis Laasfeld, Sergei Kopanchuk, Ago Rinken
Measurement of virus concentration is essential for effective virus-based transfection technologies. Here, we describe a user-friendly, image-based cell-size estimation (ICSE) assay for baculovirus quantification that relies on automated determination of cell diameters from bright-field microscopy images. In the ICSE assay, microplate-based imaging systems and our custom ICSE-Tools software enable measurement of cell morphological parameters over time. Results from the ICSE assay were in agreement with virus concentration measurements obtained using the traditional plaque assay as well as the Coulter principle-based cell-size measurement assay...
October 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/29048266/sequencing-brain-cells
#7
Janelle Weaver
Cataloging different cell types in the human brain remains a challenge for neuroscientists. Janelle Weaver speaks to researchers who think single-cell sequencing might provide a solution.
October 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/29048265/bootstrapping-biology
#8
Sarah Webb
Sarah Webb explores how DIYbio strategies are helping biologists in traditional labs do better science.
October 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28911319/convenient-and-robust-genome-wide-targeted-expression-analysis-for-biomarker-discovery
#9
(no author information available yet)
No abstract text is available yet for this article.
September 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28911318/development-and-optimization-of-a-high-titer-recombinant-lentivirus-system
#10
Austin Storck, James Ludtke, Leisha Kopp, Laura Juckem
To enable simple and effective high titer recombinant lentivirus production, we examined key parameters for the generation of lentivirus including: transfection optimization, media change, incubation time and DNA vector selection. These results illustrate the importance of optimizing transfection processes for high titer recombinant lentivirus production.
September 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28911317/generation-of-ips-derived-model-cells-for-analyses-of-hair-shaft-differentiation
#11
Takumi Kido, Tomoatsu Horigome, Minori Uda, Naoki Adachi, Yohei Hirai
Biological evaluation of hair growth/differentiation activity in vitro has been a formidable challenge, primarily due to the lack of relevant model cell systems. To solve this problem, we generated a stable model cell line in which successive differentiation via epidermal progenitors to hair components is easily inducible and traceable. Mouse induced pluripotent stem (iPS) cell-derived cells were selected to stably express a tetracycline (Tet)-inducible bone morphogenic protein-4 (BMP4) expression cassette and a luciferase reporter driven by a hair-specific keratin 31 gene (krt31) promoter (Tet-BMP4-KRT31-Luc iPS)...
September 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28911316/optimizing-t4-dna-polymerase-conditions-enhances-the-efficiency-of-one-step-sequence-and-ligation-independent-cloning
#12
Mohammad Nazrul Islam, Kyeong Won Lee, Hyung-Soon Yim, Seong Hyuk Lee, Hae Chang Jung, Jung-Hyun Lee, Jae-Yeon Jeong
Previously, we developed a one-step sequence- and ligation-independent cloning (SLIC) method that is simple, fast, and cost-effective. However, although one-step SLIC generally works well, its cloning efficiency is occasionally poor, potentially due to formation of stable secondary structures within the single-stranded DNA (ssDNA) region generated by T4 DNA polymerase during the 2.5 min treatment at room temperature. To overcome this problem, we developed a modified thermo-regulated one-step SLIC approach by testing shorter T4 DNA polymerase treatment durations (5 s-2...
September 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28911315/direct-quantification-of-protein-glycan-phosphorylation
#13
Stephanie A Ketcham, Muhammad Ashraf, Chikkathur N Madhavarao
Phosphorylation is an important post-translational modification (PTM) of proteins and a critical quality attribute for protein therapeutics, especially if it is required for protein function or sub-cellular targeting. Most current methods to quantify phosphorylation are time-consuming, indirect, or require specific instrumentation and technical skills. Here, we report the adaptation of a phosphate-specific binding dye and common laboratory instruments for quantification of relative amounts of phosphorylated glycans as well as phosphorylation of amino acid residues on the backbones of proteins...
September 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28911314/construction-of-a-combinatorial-library-of-chimeric-tumor-specific-promoters
#14
Kirill N Kashkin, Igor P Chernov, Dmitry A Didych, Eugene D Sverdlov
Gene therapy is a fast-developing field of molecular medicine. New, effective, and cancer-specific promoters are in high demand by researchers seeking to treat cancer through expression of therapeutic genes. Here, we created a combinatorial library of tumor-specific chimeric promoter modules for identifying new promoters with desired functions. The library was constructed by randomly combining promoter fragments from eight human genes involved in cell proliferation control. The pool of chimeric promoters was inserted into a lentiviral expression vector upstream of the CopGFP reporter gene, transduced into A431 cells, and enriched for active promoters by cell sorting...
September 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28911313/-letter-to-the-editor-incorrect-assignment-of-affected-nucleotides-in-footprinting-probing-experiments
#15
Sergey Belikov, Fredrik Lackmann, Lars Wieslander
Address correspondence to Sergey Belikov or Lars Wieslander, Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, SE-106 91 Stockholm, Sweden. E-mail: sergey.belikov@su.se or lars.wieslander@su.se.
September 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28911312/persistent-epigenetic-memory
#16
Amber Dance
Nematodes transmit environmental information, such as temperature or food availability, across generations. Amber Dance looks at how this is possible.
September 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28911311/plants-in-the-crispr
#17
Sarah Webb
Sarah Webb explores how researchers are using CRISPR/Cas9 to solve agricultural problems.
September 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28803546/prestige-antibodies%C3%A2-on-the-cell-atlas-from-milliporesigma
#18
(no author information available yet)
No abstract text is available yet for this article.
August 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28803545/mass-spectrometric-characterization-of-peptides-associated-with-molecules-of-the-major-histocompatibility-complex
#19
Michael R Pisano, Michael Ford
No abstract text is available yet for this article.
August 1, 2017: BioTechniques
https://www.readbyqxmd.com/read/28803544/overcoming-qrt-pcr-interference-by-select-carbon-nanotubes-in-assessments-of-gene-expression
#20
Sara T Humes, Shannon Hentschel, Candice M Lavelle, L Cody Smith, John A Lednicky, Navid B Saleh, Tara Sabo-Attwood
Nanomaterials (NMs) of various types, including carbon nanotubes (CNTs), can interfere with standard quantitative real-time PCR (qRT-PCR) assays, resulting in inaccurate gene expression measurements; however, the precise step in the qRT-PCR pipeline where this interference occurs has not been well described. Here, we investigated where in the process surface-oxidized multi-walled CNTs (oxMWNTs) inhibited qRT-PCR measurement of the expression of the housekeeping gene GAPDH and explored several strategies to minimize such inhibition...
August 1, 2017: BioTechniques
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