Various amounts of VqsMt truncated proteins were incubated with different DNA probe (Supplementary Table S2) in 25 μl of the gel shift-loading buffer (20 mM Tris-HCl, pH 7.4, 50 mM KCl, 5mM MgCl2, 10% Glycerol and 3 μg/ml sheared salmon sperm DNA). After incubation at room temperature for 20 min, the samples were analyzed by 6% polyacrylamide gel electrophoresis in 0.5X TBE (Tris/Boric Acid/EDTA) buffer at 90 V for 90 min. The gels were stained by SYBR GOLD dye and subjected to screen on a phosphor screen (BAS-IP, Fuji).
Bas ip
Manufactured by Fujifilm
The BAS-IP is a laboratory imaging system produced by Fujifilm. It is designed for the detection and analysis of radioactive samples, utilizing phosphor imaging technology. The BAS-IP provides high-resolution imaging capabilities for a range of applications in scientific research and clinical settings.
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Lab products found in correlation
3 protocols using bas ip
1
Protein-DNA Binding Assay Protocol
2
Northern Blot Analysis of RNA
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Northern blot analysis was performed as described previously (21 (link)). Total RNA was fractionated on a 6% polyacrylamide gel containing 7 M urea, and transferred to a Hybond-XL membrane (GE Healthcare). Antisense oligonucleotides were 5′ end labeled with [γ-32P] ATP (PerkinElmer Life Sciences) by T4 polynucleotide kinase (Enzynomics) and used as probes for hybridization. Hybridization was carried out in Rapid-Hyb buffer (GE Healthcare), and samples were incubated at 40°C overnight (for BC200 RNA) or at 42°C for 1 h (for the 5S RNA). The membrane was washed twice (20 min each time) at 25°C in 2× SSC buffer (20 mM sodium phosphate, pH 7.4, 0.3 M NaCl, 2 mM EDTA) containing 0.1% SDS, and twice (20 min each time) in 0.2× SSC buffer containing 0.1% SDS. The membrane was exposed to an imaging plate (Fuji BAS-IP), and the results were analyzed on a phospho-image analyzer (Fuji FLA-7000).
3
Radiolabeling of DNA Probes
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DNA probes were PCR-amplified using primers listed in Supplementary Table S1, then radiolabeled with T4 polynucleotide kinase (NEB) and [γ-32P]ATP (Perkin-Elmer). The radioactive probe (2 ng) was mixed with various amounts of the RhpR or RhpRpsph protein in 20 μl of gel shift buffer (10-mM Tris–HCl, pH 7.4, 50-mM KCl, 5-mM MgCl2, 10% glycerol, 3-μg/ml sheared salmon sperm DNA). The PSPTO_1489 promoter DNA was used for the negative control. After incubation at room temperature for 20 min, the samples were analyzed by 8% polyacrylamide gel electrophoresis (100 V for prerun, 85 V for 45 min for sample separation). The gels were dried and subjected to autoradiography on a phosphor screen (BAS-IP; Fuji). The assay was repeated at least three times with similar results.
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