RNA isolation and semi-quantitative RT-PCR analysis were performed as previously described (18 (link)). The primer, number of PCR cycles and the annealing temperature were as follows: Myosin light chain kinase (MLCK), 5′-CAACAGAGAAGACGGTGACCA-3′ (forward) and 5′-TCACAAGGCTGAAAGTCCCC-3′ (reverse), 32 cycles, 58°C; β-actin, 5′-CTTCTACAATGAGCTGCGTG-3′ (forward) and 5′-TCATGAGGTAGTCAGTCAGG-3′ (reverse), 21 cycles, 58°C. PCR products were electrophoresed on 2% agarose gels stained with ethidium bromide and visualized with a UV illuminator (FAS-V; NIPPON Genetics co., Ltd.) and analyzed by Image J 1.53e with Java 1.8.0 _172 (National Institutes of Health).
Fas 5
Manufactured by Nippon Genetics
The FAS-V is a compact and reliable flow cytometer designed for efficient cell analysis. It utilizes multiple laser excitation sources and sophisticated detector systems to enable high-performance fluorescence-activated cell sorting. The FAS-V is a versatile instrument suitable for a wide range of applications in life science research and clinical diagnostics.
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Lab products found in correlation
2 protocols using fas 5
1
Semi-quantitative RT-PCR Analysis of MLCK
2
Relaxation Assay for DNA Topoisomerase
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The relaxation assay was performed as described previously (Onoda et al., 2014; Kawano et al., 2016) . Purified enzyme and supercoiled pUC18 DNA were mixed in 10 μl of relaxation buffer (50 mM Tris-HCl (pH 8.0), 120 mM KCl, 10 mM MgCl 2 , 0.5 mM EDTA, 1 mM dithiothreitol, 0.5 mM ATP and 30 μg/ml bovine serum albumin) and incubated at 30 °C for 30 min. The reaction was stopped by adding SDS and proteinase K (Roche). After incubation at 55 °C for 1 h, the samples were subjected to 1% agarose gel electrophoresis. DNA was detected by staining with GelRed Nucleic Acid Gel Stain (Biotium). Images were captured using a Medi-Cap USB170 (MEDI CAPTURE) equipped with a CCD camera or FAS-V (NIPPON Genetics).
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