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TaKaRa ethylenediaminetetraacetic acid
Ethylenediaminetetraacetic Acid, supplied by TaKaRa, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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ethylenediaminetetraacetic acid - by Bioz Stars, 2020-04
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Histopathology:

Article Title: Human telomerase reverse transcriptase and glucose-regulated protein 78 increase the life span of articular chondrocytes and their repair potential
Article Snippet: Subsequently, the femur samples were fixed in 10% buffered formalin for 7 d. Each specimen was decalcified with 10% EDTA in distilled water (pH 7.4) for 3 weeks, and then embedded in paraffin, cut into 6-μm-thick sagittal sections, deparaffinized, and stained with safranin O (Cartilage Staining Kit, Takara, Shiga, Japan). .. The histopathology of the OA cartilage samples (n = 24) were analyzed according to standard grading and staging of OA cartilage histopathology [ ].

Centrifugation:

Article Title: Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1 2
Article Snippet: Cell debris was removed by high-speed centrifugation and the supernatant was mixed with Talon metal affinity resins (Clontech, Mountain View, CA). .. The eluted protein was dialyzed against 50 mM MES (pH 6.5), 50 mM NaCl, 10% (vol/vol) glycerol, 1 mM EDTA, and 5 mM DTT and loaded onto a MonoQ anion exchange column (Clontech). rTG2 was eluted by using a gradient of 50 to 450 mM NaCl in MES buffer.

Article Title: A Novel Glycosylphosphatidylinositol-Anchored Glycoside Hydrolase from Ustilago esculenta Functions in ?-1,3-Glucan Degradation
Article Snippet: .. The resulting pellet was washed five times with 100 mM sodium acetate (pH 5.5) containing 1 mM EDTA and treated with a mixture of Yatalase (5.4 mg, 30 U; TaKaRa Bio) and lysing enzyme (3 mg; Sigma-Aldrich) in 1.2 ml of 10 mM sodium phosphate (pH 6.0) containing 150 mM NaCl at 37°C for 2 h. The supernatant (100 μl) obtained after centrifugation for 5 min at 22,000 × g at 4°C was subjected to cold acetone precipitation. ..

Article Title: Non-proteolytic functions of calpain-3 in sarcoplasmic reticulum in skeletal muscles
Article Snippet: After centrifugation, the supernatant was incubated with either anti-calpain-3 (pIS2C) or peptide-absorbed anti-pIS2C antibody at 4°C for 1 hr. .. For the calpain-3 proteolysis assay, skeletal muscle lysates, microsomal fractions, or COS7 cell lysates were suspended in TED buffer (10 mM Tris/Cl[pH7.5], 1 mM EDTA, 1 mM DTT, 1 mM PMSF, 0.1 μM leupeptin, 40 μM bestatin, 1.5 μM aprotinin, and 0.7 μM calpastatin (Takara)), and incubated at 30°C for 30 min in the presence of 10 mM EDTA, 5 mM CaCl2, or 150 mM NaCl.

Article Title: Complex Regulation of the Bacillus subtilis Aconitase Gene
Article Snippet: Cell debris was removed by centrifugation at 20,000 × g for 20 min at 4°C. .. The suspension was centrifuged at 20,000 × g at 4°C for 15 min, and the supernatant fluids were dialyzed against 1 liter of sonication buffer without dithiothreitol or disodium EDTA for 2 h at 4°C, with buffer replacement after 1 h. The dialyzed lysates were mixed with 2 ml of Talon (Clontech) metal (Co+ ) affinity resin that had been equilibrated with buffer I (20 mM Tris-HCl [pH 8.0], 5 mM β-mercaptoethanol, 10% glycerol, 1 mM phenylmethylsulfonyl fluoride, 2 μM pepstatin A, and 0.1% Igepal) supplemented with 150 mM KCl and incubated for 20 min at 4°C with tumbling.

Amplification:

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: .. A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara). .. The reactions were stopped by adding 200 μl of ice-cold stop solution (0.4 M sodium acetate, 2.5 mM EDTA), and the DNA products were purified by phenol extraction and ethanol precipitation.

Filtration:

Article Title: A Novel Glycosylphosphatidylinositol-Anchored Glycoside Hydrolase from Ustilago esculenta Functions in ?-1,3-Glucan Degradation
Article Snippet: The culture filtrate (40 ml) obtained following the filtration of cells through cheesecloth was concentrated to 500 μl by ultrafiltration and used as the extracellular fraction. .. The resulting pellet was washed five times with 100 mM sodium acetate (pH 5.5) containing 1 mM EDTA and treated with a mixture of Yatalase (5.4 mg, 30 U; TaKaRa Bio) and lysing enzyme (3 mg; Sigma-Aldrich) in 1.2 ml of 10 mM sodium phosphate (pH 6.0) containing 150 mM NaCl at 37°C for 2 h. The supernatant (100 μl) obtained after centrifugation for 5 min at 22,000 × g at 4°C was subjected to cold acetone precipitation.

Electrophoresis:

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara). .. The digested DNA fragments were separated by electrophoresis on 6% polyacrylamide-8 M urea gels alongside sequencing ladders that had been generated with the same 32 P-labeled PerR_FP_R primer used to amplify DNA fragments for the DNase I digestion.

Article Title: Transcriptional Coactivator PC4 Stimulates Promoter Escape and Facilitates Transcriptional Synergy by GAL4-VP16
Article Snippet: For 390- and 20-nucleotide (nt) transcripts, in vitro transcription reaction mixtures (25 μl) contained 50 ng of negatively supercoiled pG5HMC2AT or its derivative, 12 mM HEPES-KOH (pH 7.9), 6% glycerol, 60 mM KCl, 0.6 mM EDTA, 8 mM MgCl2 , 5 mM dithiothreitol (DTT), 20 U of RNase inhibitor (TaKaRa), 0.2 mM ATP, 0.2 mM UTP, 0.1 mM 3′- o -methyl GTP, 12.5 μM CTP, 10 μCi of [α-32 P]CTP, 20 ng of TFIIA, 10 ng of TFIIB, 1 μl of FLAG-tagged TFIID (which corresponds to ∼0.1 ng of TATA binding protein [TBP]), 10 ng of TFIIE, 20 ng of TFIIF, 20 ng of recombinant TFIIH, and 100 ng of RNAPII. .. Transcription reactions for the initiation product were performed as described previously , and the derived pppApC was treated with calf intestinal phosphatase to form ApC before electrophoresis ( ).

Incubation:

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: .. A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara). .. The reactions were stopped by adding 200 μl of ice-cold stop solution (0.4 M sodium acetate, 2.5 mM EDTA), and the DNA products were purified by phenol extraction and ethanol precipitation.

Article Title: Rad52 prevents excessive replication fork reversal and protects from nascent strand degradation
Article Snippet: .. The reaction mixtures were incubated at 37 °C for 5 min, mixed with 1 μl of 10× Orange-G loading dye, and the free DNA was separated from the protein–DNA complexes using 0.8% Agarose (Research Products International) gel in TBE buffer (90 mM Tris [pH 8.0], 64.6 mM boric acid and 2 mM EDTA) for 2 h at 50 V using a Mupid-EX apparatus (TAKARA). .. The resolved species were visualised using a Chemi-doc (Bio-Rad) by exciting and monitoring Cy5 fluorescence.

Article Title: Non-proteolytic functions of calpain-3 in sarcoplasmic reticulum in skeletal muscles
Article Snippet: .. For the calpain-3 proteolysis assay, skeletal muscle lysates, microsomal fractions, or COS7 cell lysates were suspended in TED buffer (10 mM Tris/Cl[pH7.5], 1 mM EDTA, 1 mM DTT, 1 mM PMSF, 0.1 μM leupeptin, 40 μM bestatin, 1.5 μM aprotinin, and 0.7 μM calpastatin (Takara)), and incubated at 30°C for 30 min in the presence of 10 mM EDTA, 5 mM CaCl2, or 150 mM NaCl. ..

Article Title: Complex Regulation of the Bacillus subtilis Aconitase Gene
Article Snippet: .. The suspension was centrifuged at 20,000 × g at 4°C for 15 min, and the supernatant fluids were dialyzed against 1 liter of sonication buffer without dithiothreitol or disodium EDTA for 2 h at 4°C, with buffer replacement after 1 h. The dialyzed lysates were mixed with 2 ml of Talon (Clontech) metal (Co+ ) affinity resin that had been equilibrated with buffer I (20 mM Tris-HCl [pH 8.0], 5 mM β-mercaptoethanol, 10% glycerol, 1 mM phenylmethylsulfonyl fluoride, 2 μM pepstatin A, and 0.1% Igepal) supplemented with 150 mM KCl and incubated for 20 min at 4°C with tumbling. .. The resin and bound His-tagged protein were collected by centrifugation at 1,000 × g for 10 min at 4°C and washed sequentially at 4°C with 15 ml each of buffer I containing 500 mM KCl and 5 mM imidazole and buffer I containing 125 mM KCl and 5 mM imidazole.

Activity Assay:

Article Title: Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1 2
Article Snippet: The eluted protein was dialyzed against 50 mM MES (pH 6.5), 50 mM NaCl, 10% (vol/vol) glycerol, 1 mM EDTA, and 5 mM DTT and loaded onto a MonoQ anion exchange column (Clontech). rTG2 was eluted by using a gradient of 50 to 450 mM NaCl in MES buffer. .. Purity of rTG2 was confirmed by SDS-PAGE, and transamidation activity was assayed using a colorimetric kit (Sigma; No. CS1070), according to the manufacturer's instructions.

Expressing:

Article Title: Single nucleosome imaging reveals loose genome chromatin networks via active RNA polymerase II
Article Snippet: Paragraph title: Biochemical fractionation of nuclei from cells expressing H2B-HaloTag ... Briefly, collected cells were suspended in nuclei isolation buffer (3.75 mM Tris-HCl [pH 7.5], 20 mM KCl, 0.5 mM EDTA, 0.05 mM spermine, 0.125 mM spermidine, 1 µg/ml Aprotinin [T010A; TaKaRa], and 0.1 mM PMSF [P7626-1G; Sigma-Aldrich]) and centrifuged at 1,936 g for 7 min at room temperature.

Article Title: Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1 2
Article Snippet: Protein expression was induced with 300 µM isopropylthio-β-galactoside (IPTG) for 24 hours at 25°C. .. The eluted protein was dialyzed against 50 mM MES (pH 6.5), 50 mM NaCl, 10% (vol/vol) glycerol, 1 mM EDTA, and 5 mM DTT and loaded onto a MonoQ anion exchange column (Clontech). rTG2 was eluted by using a gradient of 50 to 450 mM NaCl in MES buffer.

Article Title: Pentoxifylline Reduces Tumor Necrosis Factor-? and HIV-Induced Vascular Endothelial Activation
Article Snippet: For analysis of VCAM-1 and IP-10 gene expression and protein release, 4.0×104 HLMVECs between passages 3 and 6 were seeded in each well of 12-well tissue culture plates. .. After the indicated time points, the supernatant was harvested for IP-10 analysis and the cells were either removed from the plate using 5 mM EDTA in phosphate-buffered saline (PBS) for staining analysis or lysed and the RNA harvested using a NucleoSpin RNA isolation kit (Clontech Laboratories, Mountain View, CA) for reverse transcriptase polymerase chain reaction (RT-PCR).

Article Title: Complex Regulation of the Bacillus subtilis Aconitase Gene
Article Snippet: When the culture reached an optical density at 600 nm (OD600 ) of approximately 0.6, expression of CodY-His6 was induced by the addition of arabinose to a final concentration of 0.2%. .. The suspension was centrifuged at 20,000 × g at 4°C for 15 min, and the supernatant fluids were dialyzed against 1 liter of sonication buffer without dithiothreitol or disodium EDTA for 2 h at 4°C, with buffer replacement after 1 h. The dialyzed lysates were mixed with 2 ml of Talon (Clontech) metal (Co+ ) affinity resin that had been equilibrated with buffer I (20 mM Tris-HCl [pH 8.0], 5 mM β-mercaptoethanol, 10% glycerol, 1 mM phenylmethylsulfonyl fluoride, 2 μM pepstatin A, and 0.1% Igepal) supplemented with 150 mM KCl and incubated for 20 min at 4°C with tumbling.

Recombinase Polymerase Amplification:

Article Title: Rad52 prevents excessive replication fork reversal and protects from nascent strand degradation
Article Snippet: EMSAs In all, 20 nM of G1 or RF1 DNA were mixed with the indicated concentrations of RPA and RAD52 in 10 μl of standard reaction buffer, containing 10 mM Tris-Acetate [pH 7.5], 1 mM DTT, 0.1 μg/ml BSA, 150 mM NaCl and 5 mM magnesium acetate. .. The reaction mixtures were incubated at 37 °C for 5 min, mixed with 1 μl of 10× Orange-G loading dye, and the free DNA was separated from the protein–DNA complexes using 0.8% Agarose (Research Products International) gel in TBE buffer (90 mM Tris [pH 8.0], 64.6 mM boric acid and 2 mM EDTA) for 2 h at 50 V using a Mupid-EX apparatus (TAKARA).

Proteolysis Assay:

Article Title: Non-proteolytic functions of calpain-3 in sarcoplasmic reticulum in skeletal muscles
Article Snippet: .. For the calpain-3 proteolysis assay, skeletal muscle lysates, microsomal fractions, or COS7 cell lysates were suspended in TED buffer (10 mM Tris/Cl[pH7.5], 1 mM EDTA, 1 mM DTT, 1 mM PMSF, 0.1 μM leupeptin, 40 μM bestatin, 1.5 μM aprotinin, and 0.7 μM calpastatin (Takara)), and incubated at 30°C for 30 min in the presence of 10 mM EDTA, 5 mM CaCl2, or 150 mM NaCl. ..

Derivative Assay:

Article Title: Transcriptional Coactivator PC4 Stimulates Promoter Escape and Facilitates Transcriptional Synergy by GAL4-VP16
Article Snippet: For 390- and 20-nucleotide (nt) transcripts, in vitro transcription reaction mixtures (25 μl) contained 50 ng of negatively supercoiled pG5HMC2AT or its derivative, 12 mM HEPES-KOH (pH 7.9), 6% glycerol, 60 mM KCl, 0.6 mM EDTA, 8 mM MgCl2 , 5 mM dithiothreitol (DTT), 20 U of RNase inhibitor (TaKaRa), 0.2 mM ATP, 0.2 mM UTP, 0.1 mM 3′- o -methyl GTP, 12.5 μM CTP, 10 μCi of [α-32 P]CTP, 20 ng of TFIIA, 10 ng of TFIIB, 1 μl of FLAG-tagged TFIID (which corresponds to ∼0.1 ng of TATA binding protein [TBP]), 10 ng of TFIIE, 20 ng of TFIIF, 20 ng of recombinant TFIIH, and 100 ng of RNAPII. .. Transcription reactions for the initiation product were performed as described previously , and the derived pppApC was treated with calf intestinal phosphatase to form ApC before electrophoresis ( ).

Immunohistochemistry:

Article Title: Human telomerase reverse transcriptase and glucose-regulated protein 78 increase the life span of articular chondrocytes and their repair potential
Article Snippet: Subsequently, the femur samples were fixed in 10% buffered formalin for 7 d. Each specimen was decalcified with 10% EDTA in distilled water (pH 7.4) for 3 weeks, and then embedded in paraffin, cut into 6-μm-thick sagittal sections, deparaffinized, and stained with safranin O (Cartilage Staining Kit, Takara, Shiga, Japan). .. Immunohistochemical staining for type II collagen was performed as described previously [ ].

Over Expression:

Article Title: Complex Regulation of the Bacillus subtilis Aconitase Gene
Article Snippet: Paragraph title: Protein overexpression and purification. ... The suspension was centrifuged at 20,000 × g at 4°C for 15 min, and the supernatant fluids were dialyzed against 1 liter of sonication buffer without dithiothreitol or disodium EDTA for 2 h at 4°C, with buffer replacement after 1 h. The dialyzed lysates were mixed with 2 ml of Talon (Clontech) metal (Co+ ) affinity resin that had been equilibrated with buffer I (20 mM Tris-HCl [pH 8.0], 5 mM β-mercaptoethanol, 10% glycerol, 1 mM phenylmethylsulfonyl fluoride, 2 μM pepstatin A, and 0.1% Igepal) supplemented with 150 mM KCl and incubated for 20 min at 4°C with tumbling.

Immunoprecipitation:

Article Title: Non-proteolytic functions of calpain-3 in sarcoplasmic reticulum in skeletal muscles
Article Snippet: Paragraph title: Immunoprecipitation and proteolysis assay of calpain-3 ... For the calpain-3 proteolysis assay, skeletal muscle lysates, microsomal fractions, or COS7 cell lysates were suspended in TED buffer (10 mM Tris/Cl[pH7.5], 1 mM EDTA, 1 mM DTT, 1 mM PMSF, 0.1 μM leupeptin, 40 μM bestatin, 1.5 μM aprotinin, and 0.7 μM calpastatin (Takara)), and incubated at 30°C for 30 min in the presence of 10 mM EDTA, 5 mM CaCl2, or 150 mM NaCl.

Footprinting:

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: Paragraph title: DNase I footprinting assays. ... A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara).

Northern Blot:

Article Title: TRF4 Is Involved in Polyadenylation of snRNAs in Drosophila melanogaster ▿
Article Snippet: Paragraph title: RNA purification, Northern blotting, and reverse transcription-PCR (RT-PCR) analysis. ... The mixture was adjusted to 53 mM Tris-HCl (pH 7.9), 25 mM NaCl, 0.5 mM EDTA, 1 mM dithiothreitol, and 30 μg of bovine serum albumin (BSA)/ml and digested for 1 h at 37°C with 60 U of RNase H (TaKaRa Bio., Inc.).

Reverse Transcription Polymerase Chain Reaction:

Article Title: TRF4 Is Involved in Polyadenylation of snRNAs in Drosophila melanogaster ▿
Article Snippet: Paragraph title: RNA purification, Northern blotting, and reverse transcription-PCR (RT-PCR) analysis. ... The mixture was adjusted to 53 mM Tris-HCl (pH 7.9), 25 mM NaCl, 0.5 mM EDTA, 1 mM dithiothreitol, and 30 μg of bovine serum albumin (BSA)/ml and digested for 1 h at 37°C with 60 U of RNase H (TaKaRa Bio., Inc.).

Article Title: Pentoxifylline Reduces Tumor Necrosis Factor-? and HIV-Induced Vascular Endothelial Activation
Article Snippet: .. After the indicated time points, the supernatant was harvested for IP-10 analysis and the cells were either removed from the plate using 5 mM EDTA in phosphate-buffered saline (PBS) for staining analysis or lysed and the RNA harvested using a NucleoSpin RNA isolation kit (Clontech Laboratories, Mountain View, CA) for reverse transcriptase polymerase chain reaction (RT-PCR). .. Total RNA was extracted and reactions were performed in duplicate on 96-well optical reaction plates with one-step SYBR Green PCR master mix (iScript One-Step RT-PCT Kit with SYBR Green, Bio-Rad Laboratories, Hercules, CA).

Light Microscopy:

Article Title: Human telomerase reverse transcriptase and glucose-regulated protein 78 increase the life span of articular chondrocytes and their repair potential
Article Snippet: Postoperative analyses Eight and 16 weeks after implantation, rabbits were killed with an overdose of intravenous anesthesia, and then the distal parts of their femurs were harvested and observed with a light microscope. .. Subsequently, the femur samples were fixed in 10% buffered formalin for 7 d. Each specimen was decalcified with 10% EDTA in distilled water (pH 7.4) for 3 weeks, and then embedded in paraffin, cut into 6-μm-thick sagittal sections, deparaffinized, and stained with safranin O (Cartilage Staining Kit, Takara, Shiga, Japan).

Generated:

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara). .. The digested DNA fragments were separated by electrophoresis on 6% polyacrylamide-8 M urea gels alongside sequencing ladders that had been generated with the same 32 P-labeled PerR_FP_R primer used to amplify DNA fragments for the DNase I digestion.

Polymerase Chain Reaction:

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: .. A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara). .. The reactions were stopped by adding 200 μl of ice-cold stop solution (0.4 M sodium acetate, 2.5 mM EDTA), and the DNA products were purified by phenol extraction and ethanol precipitation.

Article Title: Pentoxifylline Reduces Tumor Necrosis Factor-? and HIV-Induced Vascular Endothelial Activation
Article Snippet: .. After the indicated time points, the supernatant was harvested for IP-10 analysis and the cells were either removed from the plate using 5 mM EDTA in phosphate-buffered saline (PBS) for staining analysis or lysed and the RNA harvested using a NucleoSpin RNA isolation kit (Clontech Laboratories, Mountain View, CA) for reverse transcriptase polymerase chain reaction (RT-PCR). .. Total RNA was extracted and reactions were performed in duplicate on 96-well optical reaction plates with one-step SYBR Green PCR master mix (iScript One-Step RT-PCT Kit with SYBR Green, Bio-Rad Laboratories, Hercules, CA).

Article Title: DNA Barcoding studies on Thrips in India: Cryptic species and Species complexes
Article Snippet: .. The PCR reaction was set in a 50 µl total volume containing 20 Picomoles of each primer, 20 mM Tris-HCl (pH 8.0), 100 mM KCl, 0.1 mM EDTA,1 mM DTT, 1.8 mM MgCl2, 0.25 mM of each dNTP, and 1U of Taq polymerase (Takara BIO Inc., Japan) with the following cycling parameters: 5 min at 94 °C; followed by 40 cycles of 30 s at 94 °C, 40 s at 49 °C, 1 min at 72 °C and final extension for 5 min at 72 °C. ..

Sonication:

Article Title: Complex Regulation of the Bacillus subtilis Aconitase Gene
Article Snippet: .. The suspension was centrifuged at 20,000 × g at 4°C for 15 min, and the supernatant fluids were dialyzed against 1 liter of sonication buffer without dithiothreitol or disodium EDTA for 2 h at 4°C, with buffer replacement after 1 h. The dialyzed lysates were mixed with 2 ml of Talon (Clontech) metal (Co+ ) affinity resin that had been equilibrated with buffer I (20 mM Tris-HCl [pH 8.0], 5 mM β-mercaptoethanol, 10% glycerol, 1 mM phenylmethylsulfonyl fluoride, 2 μM pepstatin A, and 0.1% Igepal) supplemented with 150 mM KCl and incubated for 20 min at 4°C with tumbling. .. The resin and bound His-tagged protein were collected by centrifugation at 1,000 × g for 10 min at 4°C and washed sequentially at 4°C with 15 ml each of buffer I containing 500 mM KCl and 5 mM imidazole and buffer I containing 125 mM KCl and 5 mM imidazole.

Recombinant:

Article Title: A Novel Glycosylphosphatidylinositol-Anchored Glycoside Hydrolase from Ustilago esculenta Functions in ?-1,3-Glucan Degradation
Article Snippet: Paragraph title: Fractionation of recombinant Lam16A. ... The resulting pellet was washed five times with 100 mM sodium acetate (pH 5.5) containing 1 mM EDTA and treated with a mixture of Yatalase (5.4 mg, 30 U; TaKaRa Bio) and lysing enzyme (3 mg; Sigma-Aldrich) in 1.2 ml of 10 mM sodium phosphate (pH 6.0) containing 150 mM NaCl at 37°C for 2 h. The supernatant (100 μl) obtained after centrifugation for 5 min at 22,000 × g at 4°C was subjected to cold acetone precipitation.

Article Title: Transcriptional Coactivator PC4 Stimulates Promoter Escape and Facilitates Transcriptional Synergy by GAL4-VP16
Article Snippet: .. For 390- and 20-nucleotide (nt) transcripts, in vitro transcription reaction mixtures (25 μl) contained 50 ng of negatively supercoiled pG5HMC2AT or its derivative, 12 mM HEPES-KOH (pH 7.9), 6% glycerol, 60 mM KCl, 0.6 mM EDTA, 8 mM MgCl2 , 5 mM dithiothreitol (DTT), 20 U of RNase inhibitor (TaKaRa), 0.2 mM ATP, 0.2 mM UTP, 0.1 mM 3′- o -methyl GTP, 12.5 μM CTP, 10 μCi of [α-32 P]CTP, 20 ng of TFIIA, 10 ng of TFIIB, 1 μl of FLAG-tagged TFIID (which corresponds to ∼0.1 ng of TATA binding protein [TBP]), 10 ng of TFIIE, 20 ng of TFIIF, 20 ng of recombinant TFIIH, and 100 ng of RNAPII. ..

Staining:

Article Title: Human telomerase reverse transcriptase and glucose-regulated protein 78 increase the life span of articular chondrocytes and their repair potential
Article Snippet: .. Subsequently, the femur samples were fixed in 10% buffered formalin for 7 d. Each specimen was decalcified with 10% EDTA in distilled water (pH 7.4) for 3 weeks, and then embedded in paraffin, cut into 6-μm-thick sagittal sections, deparaffinized, and stained with safranin O (Cartilage Staining Kit, Takara, Shiga, Japan). .. The histopathology of the OA cartilage samples (n = 24) were analyzed according to standard grading and staging of OA cartilage histopathology [ ].

Article Title: Pentoxifylline Reduces Tumor Necrosis Factor-? and HIV-Induced Vascular Endothelial Activation
Article Snippet: .. After the indicated time points, the supernatant was harvested for IP-10 analysis and the cells were either removed from the plate using 5 mM EDTA in phosphate-buffered saline (PBS) for staining analysis or lysed and the RNA harvested using a NucleoSpin RNA isolation kit (Clontech Laboratories, Mountain View, CA) for reverse transcriptase polymerase chain reaction (RT-PCR). .. Total RNA was extracted and reactions were performed in duplicate on 96-well optical reaction plates with one-step SYBR Green PCR master mix (iScript One-Step RT-PCT Kit with SYBR Green, Bio-Rad Laboratories, Hercules, CA).

DNA Extraction:

Article Title: DNA Barcoding studies on Thrips in India: Cryptic species and Species complexes
Article Snippet: Paragraph title: DNA extraction, Polymerase Chain Reaction and Sequencing ... The PCR reaction was set in a 50 µl total volume containing 20 Picomoles of each primer, 20 mM Tris-HCl (pH 8.0), 100 mM KCl, 0.1 mM EDTA,1 mM DTT, 1.8 mM MgCl2, 0.25 mM of each dNTP, and 1U of Taq polymerase (Takara BIO Inc., Japan) with the following cycling parameters: 5 min at 94 °C; followed by 40 cycles of 30 s at 94 °C, 40 s at 49 °C, 1 min at 72 °C and final extension for 5 min at 72 °C.

Fluorescence:

Article Title: Rad52 prevents excessive replication fork reversal and protects from nascent strand degradation
Article Snippet: The reaction mixtures were incubated at 37 °C for 5 min, mixed with 1 μl of 10× Orange-G loading dye, and the free DNA was separated from the protein–DNA complexes using 0.8% Agarose (Research Products International) gel in TBE buffer (90 mM Tris [pH 8.0], 64.6 mM boric acid and 2 mM EDTA) for 2 h at 50 V using a Mupid-EX apparatus (TAKARA). .. The resolved species were visualised using a Chemi-doc (Bio-Rad) by exciting and monitoring Cy5 fluorescence.

Isolation:

Article Title: Single nucleosome imaging reveals loose genome chromatin networks via active RNA polymerase II
Article Snippet: .. Briefly, collected cells were suspended in nuclei isolation buffer (3.75 mM Tris-HCl [pH 7.5], 20 mM KCl, 0.5 mM EDTA, 0.05 mM spermine, 0.125 mM spermidine, 1 µg/ml Aprotinin [T010A; TaKaRa], and 0.1 mM PMSF [P7626-1G; Sigma-Aldrich]) and centrifuged at 1,936 g for 7 min at room temperature. .. The cell pellets were resuspended in nuclei isolation buffer and again centrifuged at 1,936 g for 7 min at room temperature.

Article Title: TRF4 Is Involved in Polyadenylation of snRNAs in Drosophila melanogaster ▿
Article Snippet: Poly(A)+ RNA was purified from total RNA using PolyATract mRNA isolation kit according to the manufacturer's protocols (Promega, Madison, WI). .. The mixture was adjusted to 53 mM Tris-HCl (pH 7.9), 25 mM NaCl, 0.5 mM EDTA, 1 mM dithiothreitol, and 30 μg of bovine serum albumin (BSA)/ml and digested for 1 h at 37°C with 60 U of RNase H (TaKaRa Bio., Inc.).

Article Title: Pentoxifylline Reduces Tumor Necrosis Factor-? and HIV-Induced Vascular Endothelial Activation
Article Snippet: .. After the indicated time points, the supernatant was harvested for IP-10 analysis and the cells were either removed from the plate using 5 mM EDTA in phosphate-buffered saline (PBS) for staining analysis or lysed and the RNA harvested using a NucleoSpin RNA isolation kit (Clontech Laboratories, Mountain View, CA) for reverse transcriptase polymerase chain reaction (RT-PCR). .. Total RNA was extracted and reactions were performed in duplicate on 96-well optical reaction plates with one-step SYBR Green PCR master mix (iScript One-Step RT-PCT Kit with SYBR Green, Bio-Rad Laboratories, Hercules, CA).

Purification:

Article Title: TRF4 Is Involved in Polyadenylation of snRNAs in Drosophila melanogaster ▿
Article Snippet: Paragraph title: RNA purification, Northern blotting, and reverse transcription-PCR (RT-PCR) analysis. ... The mixture was adjusted to 53 mM Tris-HCl (pH 7.9), 25 mM NaCl, 0.5 mM EDTA, 1 mM dithiothreitol, and 30 μg of bovine serum albumin (BSA)/ml and digested for 1 h at 37°C with 60 U of RNase H (TaKaRa Bio., Inc.).

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: .. A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara). .. The reactions were stopped by adding 200 μl of ice-cold stop solution (0.4 M sodium acetate, 2.5 mM EDTA), and the DNA products were purified by phenol extraction and ethanol precipitation.

Article Title: Transcriptional Coactivator PC4 Stimulates Promoter Escape and Facilitates Transcriptional Synergy by GAL4-VP16
Article Snippet: Paragraph title: Purification of transcription factors and in vitro transcription assays. ... For 390- and 20-nucleotide (nt) transcripts, in vitro transcription reaction mixtures (25 μl) contained 50 ng of negatively supercoiled pG5HMC2AT or its derivative, 12 mM HEPES-KOH (pH 7.9), 6% glycerol, 60 mM KCl, 0.6 mM EDTA, 8 mM MgCl2 , 5 mM dithiothreitol (DTT), 20 U of RNase inhibitor (TaKaRa), 0.2 mM ATP, 0.2 mM UTP, 0.1 mM 3′- o -methyl GTP, 12.5 μM CTP, 10 μCi of [α-32 P]CTP, 20 ng of TFIIA, 10 ng of TFIIB, 1 μl of FLAG-tagged TFIID (which corresponds to ∼0.1 ng of TATA binding protein [TBP]), 10 ng of TFIIE, 20 ng of TFIIF, 20 ng of recombinant TFIIH, and 100 ng of RNAPII.

Article Title: Complex Regulation of the Bacillus subtilis Aconitase Gene
Article Snippet: Paragraph title: Protein overexpression and purification. ... The suspension was centrifuged at 20,000 × g at 4°C for 15 min, and the supernatant fluids were dialyzed against 1 liter of sonication buffer without dithiothreitol or disodium EDTA for 2 h at 4°C, with buffer replacement after 1 h. The dialyzed lysates were mixed with 2 ml of Talon (Clontech) metal (Co+ ) affinity resin that had been equilibrated with buffer I (20 mM Tris-HCl [pH 8.0], 5 mM β-mercaptoethanol, 10% glycerol, 1 mM phenylmethylsulfonyl fluoride, 2 μM pepstatin A, and 0.1% Igepal) supplemented with 150 mM KCl and incubated for 20 min at 4°C with tumbling.

Article Title: DNA Barcoding studies on Thrips in India: Cryptic species and Species complexes
Article Snippet: The PCR reaction was set in a 50 µl total volume containing 20 Picomoles of each primer, 20 mM Tris-HCl (pH 8.0), 100 mM KCl, 0.1 mM EDTA,1 mM DTT, 1.8 mM MgCl2, 0.25 mM of each dNTP, and 1U of Taq polymerase (Takara BIO Inc., Japan) with the following cycling parameters: 5 min at 94 °C; followed by 40 cycles of 30 s at 94 °C, 40 s at 49 °C, 1 min at 72 °C and final extension for 5 min at 72 °C. .. The PCR amplified products were purified using the QIAquick Gel Extraction Kit (Qiagen, Valencia, CA) following the manufacturer’s protocols.

Sequencing:

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara). .. The digested DNA fragments were separated by electrophoresis on 6% polyacrylamide-8 M urea gels alongside sequencing ladders that had been generated with the same 32 P-labeled PerR_FP_R primer used to amplify DNA fragments for the DNase I digestion.

Article Title: DNA Barcoding studies on Thrips in India: Cryptic species and Species complexes
Article Snippet: Paragraph title: DNA extraction, Polymerase Chain Reaction and Sequencing ... The PCR reaction was set in a 50 µl total volume containing 20 Picomoles of each primer, 20 mM Tris-HCl (pH 8.0), 100 mM KCl, 0.1 mM EDTA,1 mM DTT, 1.8 mM MgCl2, 0.25 mM of each dNTP, and 1U of Taq polymerase (Takara BIO Inc., Japan) with the following cycling parameters: 5 min at 94 °C; followed by 40 cycles of 30 s at 94 °C, 40 s at 49 °C, 1 min at 72 °C and final extension for 5 min at 72 °C.

Lysis:

Article Title: Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1 2
Article Snippet: Cell pellets were lysed by sonication at 0°C in lysis buffer [50 mM Na2 HPO4 (pH 7.5), 400 mM NaCl, 5 mM benzamidine, and 5 mM 2-mercaptoethanol] containing 50 µM GTP, 50 µMadenosine-5′-triphosphate (ATP), and 50 µg/ml phenylmethanesulfonyl fluoride (PMSF). .. The eluted protein was dialyzed against 50 mM MES (pH 6.5), 50 mM NaCl, 10% (vol/vol) glycerol, 1 mM EDTA, and 5 mM DTT and loaded onto a MonoQ anion exchange column (Clontech). rTG2 was eluted by using a gradient of 50 to 450 mM NaCl in MES buffer.

SDS Page:

Article Title: Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1Extracellular Tissue Transglutaminase Activates Noncanonical NF-?B Signaling and Promotes Metastasis in Ovarian Cancer 1 2
Article Snippet: The eluted protein was dialyzed against 50 mM MES (pH 6.5), 50 mM NaCl, 10% (vol/vol) glycerol, 1 mM EDTA, and 5 mM DTT and loaded onto a MonoQ anion exchange column (Clontech). rTG2 was eluted by using a gradient of 50 to 450 mM NaCl in MES buffer. .. Purity of rTG2 was confirmed by SDS-PAGE, and transamidation activity was assayed using a colorimetric kit (Sigma; No. CS1070), according to the manufacturer's instructions.

Binding Assay:

Article Title: Transcriptional Coactivator PC4 Stimulates Promoter Escape and Facilitates Transcriptional Synergy by GAL4-VP16
Article Snippet: .. For 390- and 20-nucleotide (nt) transcripts, in vitro transcription reaction mixtures (25 μl) contained 50 ng of negatively supercoiled pG5HMC2AT or its derivative, 12 mM HEPES-KOH (pH 7.9), 6% glycerol, 60 mM KCl, 0.6 mM EDTA, 8 mM MgCl2 , 5 mM dithiothreitol (DTT), 20 U of RNase inhibitor (TaKaRa), 0.2 mM ATP, 0.2 mM UTP, 0.1 mM 3′- o -methyl GTP, 12.5 μM CTP, 10 μCi of [α-32 P]CTP, 20 ng of TFIIA, 10 ng of TFIIB, 1 μl of FLAG-tagged TFIID (which corresponds to ∼0.1 ng of TATA binding protein [TBP]), 10 ng of TFIIE, 20 ng of TFIIF, 20 ng of recombinant TFIIH, and 100 ng of RNAPII. ..

Article Title: Non-proteolytic functions of calpain-3 in sarcoplasmic reticulum in skeletal muscles
Article Snippet: To minimize non-specific binding, the microsomal fraction was incubated with protein-G-Sepharose 4FF (GE) at 4°C for 1 hr. .. For the calpain-3 proteolysis assay, skeletal muscle lysates, microsomal fractions, or COS7 cell lysates were suspended in TED buffer (10 mM Tris/Cl[pH7.5], 1 mM EDTA, 1 mM DTT, 1 mM PMSF, 0.1 μM leupeptin, 40 μM bestatin, 1.5 μM aprotinin, and 0.7 μM calpastatin (Takara)), and incubated at 30°C for 30 min in the presence of 10 mM EDTA, 5 mM CaCl2, or 150 mM NaCl.

Agarose Gel Electrophoresis:

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: .. A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara). .. The reactions were stopped by adding 200 μl of ice-cold stop solution (0.4 M sodium acetate, 2.5 mM EDTA), and the DNA products were purified by phenol extraction and ethanol precipitation.

In Vitro:

Article Title: Pentoxifylline Reduces Tumor Necrosis Factor-? and HIV-Induced Vascular Endothelial Activation
Article Snippet: Paragraph title: In vitro assays ... After the indicated time points, the supernatant was harvested for IP-10 analysis and the cells were either removed from the plate using 5 mM EDTA in phosphate-buffered saline (PBS) for staining analysis or lysed and the RNA harvested using a NucleoSpin RNA isolation kit (Clontech Laboratories, Mountain View, CA) for reverse transcriptase polymerase chain reaction (RT-PCR).

Article Title: Transcriptional Coactivator PC4 Stimulates Promoter Escape and Facilitates Transcriptional Synergy by GAL4-VP16
Article Snippet: .. For 390- and 20-nucleotide (nt) transcripts, in vitro transcription reaction mixtures (25 μl) contained 50 ng of negatively supercoiled pG5HMC2AT or its derivative, 12 mM HEPES-KOH (pH 7.9), 6% glycerol, 60 mM KCl, 0.6 mM EDTA, 8 mM MgCl2 , 5 mM dithiothreitol (DTT), 20 U of RNase inhibitor (TaKaRa), 0.2 mM ATP, 0.2 mM UTP, 0.1 mM 3′- o -methyl GTP, 12.5 μM CTP, 10 μCi of [α-32 P]CTP, 20 ng of TFIIA, 10 ng of TFIIB, 1 μl of FLAG-tagged TFIID (which corresponds to ∼0.1 ng of TATA binding protein [TBP]), 10 ng of TFIIE, 20 ng of TFIIF, 20 ng of recombinant TFIIH, and 100 ng of RNAPII. ..

Ethanol Precipitation:

Article Title: Regulation of perR Expression by Iron and PerR in Campylobacter jejuni ▿ ▿ †
Article Snippet: A DNA region containing the perR promoter was PCR amplified with a 32 P-labeled primer perR_FP_R and a nonradiolabeled primer perR_FP_F, and the PCR product was purified from an agarose gel with a Wizard SV Gel and PCR Clean-Up System (Promega). rPerR was incubated with the 32 P-labeled perR promoter at 37°C for 15 min in 40 μl of the DNA-binding buffer [20 mM HEPES (pH 7.6), 1 mM EDTA, 10 mM (NH4 )2 SO4 , 5 mM DTT, 0.2% Tween 20, 30 mM KCl, 10 mM MgCl2 , and 0.1 μg of poly(dI-dC)], and the reaction mixture was treated with 0.1 U of DNase I (Takara). .. The reactions were stopped by adding 200 μl of ice-cold stop solution (0.4 M sodium acetate, 2.5 mM EDTA), and the DNA products were purified by phenol extraction and ethanol precipitation.

Concentration Assay:

Article Title: Complex Regulation of the Bacillus subtilis Aconitase Gene
Article Snippet: When the culture reached an optical density at 600 nm (OD600 ) of approximately 0.6, expression of CodY-His6 was induced by the addition of arabinose to a final concentration of 0.2%. .. The suspension was centrifuged at 20,000 × g at 4°C for 15 min, and the supernatant fluids were dialyzed against 1 liter of sonication buffer without dithiothreitol or disodium EDTA for 2 h at 4°C, with buffer replacement after 1 h. The dialyzed lysates were mixed with 2 ml of Talon (Clontech) metal (Co+ ) affinity resin that had been equilibrated with buffer I (20 mM Tris-HCl [pH 8.0], 5 mM β-mercaptoethanol, 10% glycerol, 1 mM phenylmethylsulfonyl fluoride, 2 μM pepstatin A, and 0.1% Igepal) supplemented with 150 mM KCl and incubated for 20 min at 4°C with tumbling.

Fractionation:

Article Title: Single nucleosome imaging reveals loose genome chromatin networks via active RNA polymerase II
Article Snippet: Paragraph title: Biochemical fractionation of nuclei from cells expressing H2B-HaloTag ... Briefly, collected cells were suspended in nuclei isolation buffer (3.75 mM Tris-HCl [pH 7.5], 20 mM KCl, 0.5 mM EDTA, 0.05 mM spermine, 0.125 mM spermidine, 1 µg/ml Aprotinin [T010A; TaKaRa], and 0.1 mM PMSF [P7626-1G; Sigma-Aldrich]) and centrifuged at 1,936 g for 7 min at room temperature.

Article Title: A Novel Glycosylphosphatidylinositol-Anchored Glycoside Hydrolase from Ustilago esculenta Functions in ?-1,3-Glucan Degradation
Article Snippet: Paragraph title: Fractionation of recombinant Lam16A. ... The resulting pellet was washed five times with 100 mM sodium acetate (pH 5.5) containing 1 mM EDTA and treated with a mixture of Yatalase (5.4 mg, 30 U; TaKaRa Bio) and lysing enzyme (3 mg; Sigma-Aldrich) in 1.2 ml of 10 mM sodium phosphate (pH 6.0) containing 150 mM NaCl at 37°C for 2 h. The supernatant (100 μl) obtained after centrifugation for 5 min at 22,000 × g at 4°C was subjected to cold acetone precipitation.

Gel Extraction:

Article Title: DNA Barcoding studies on Thrips in India: Cryptic species and Species complexes
Article Snippet: The PCR reaction was set in a 50 µl total volume containing 20 Picomoles of each primer, 20 mM Tris-HCl (pH 8.0), 100 mM KCl, 0.1 mM EDTA,1 mM DTT, 1.8 mM MgCl2, 0.25 mM of each dNTP, and 1U of Taq polymerase (Takara BIO Inc., Japan) with the following cycling parameters: 5 min at 94 °C; followed by 40 cycles of 30 s at 94 °C, 40 s at 49 °C, 1 min at 72 °C and final extension for 5 min at 72 °C. .. The PCR amplified products were purified using the QIAquick Gel Extraction Kit (Qiagen, Valencia, CA) following the manufacturer’s protocols.

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  • 86
    TaKaRa bal31 buffer
    Detection of terminal fragments of minichromosomes V4 ( A and B ) and Y32 ( C and D ). Chromosomal DNA was first treated with <t>Bal31</t> for various periods and then with HincII (A and B) or HindIII (C and D). Hybridizing bands were detected with an 800 bp 3′-region of Zepp or with the telomeric repeats (pCHt1) as a probe (B and D). Arrowheads indicate hybridizing bands derived from chromosome termini. Lanes 1–6, Bal31 treatment for 0, 30, 60, 120, 180 and 300 s, respectively.
    Bal31 Buffer, supplied by TaKaRa, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/bal31 buffer/product/TaKaRa
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    bal31 buffer - by Bioz Stars, 2020-04
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    edta  (TaKaRa)
    94
    TaKaRa edta
    Detection of terminal fragments of minichromosomes V4 ( A and B ) and Y32 ( C and D ). Chromosomal DNA was first treated with <t>Bal31</t> for various periods and then with HincII (A and B) or HindIII (C and D). Hybridizing bands were detected with an 800 bp 3′-region of Zepp or with the telomeric repeats (pCHt1) as a probe (B and D). Arrowheads indicate hybridizing bands derived from chromosome termini. Lanes 1–6, Bal31 treatment for 0, 30, 60, 120, 180 and 300 s, respectively.
    Edta, supplied by TaKaRa, used in various techniques. Bioz Stars score: 94/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/edta/product/TaKaRa
    Average 94 stars, based on 17 article reviews
    Price from $9.99 to $1999.99
    edta - by Bioz Stars, 2020-04
    94/100 stars
      Buy from Supplier

    99
    TaKaRa proteoguard edta free protease inhibitor cocktail
    Detection of terminal fragments of minichromosomes V4 ( A and B ) and Y32 ( C and D ). Chromosomal DNA was first treated with <t>Bal31</t> for various periods and then with HincII (A and B) or HindIII (C and D). Hybridizing bands were detected with an 800 bp 3′-region of Zepp or with the telomeric repeats (pCHt1) as a probe (B and D). Arrowheads indicate hybridizing bands derived from chromosome termini. Lanes 1–6, Bal31 treatment for 0, 30, 60, 120, 180 and 300 s, respectively.
    Proteoguard Edta Free Protease Inhibitor Cocktail, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/proteoguard edta free protease inhibitor cocktail/product/TaKaRa
    Average 99 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    proteoguard edta free protease inhibitor cocktail - by Bioz Stars, 2020-04
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    Detection of terminal fragments of minichromosomes V4 ( A and B ) and Y32 ( C and D ). Chromosomal DNA was first treated with Bal31 for various periods and then with HincII (A and B) or HindIII (C and D). Hybridizing bands were detected with an 800 bp 3′-region of Zepp or with the telomeric repeats (pCHt1) as a probe (B and D). Arrowheads indicate hybridizing bands derived from chromosome termini. Lanes 1–6, Bal31 treatment for 0, 30, 60, 120, 180 and 300 s, respectively.

    Journal: Nucleic Acids Research

    Article Title: Retrotransposon-mediated restoration of Chlorella telomeres: accumulation of Zepp retrotransposons at termini of newly formed minichromosomes

    doi:

    Figure Lengend Snippet: Detection of terminal fragments of minichromosomes V4 ( A and B ) and Y32 ( C and D ). Chromosomal DNA was first treated with Bal31 for various periods and then with HincII (A and B) or HindIII (C and D). Hybridizing bands were detected with an 800 bp 3′-region of Zepp or with the telomeric repeats (pCHt1) as a probe (B and D). Arrowheads indicate hybridizing bands derived from chromosome termini. Lanes 1–6, Bal31 treatment for 0, 30, 60, 120, 180 and 300 s, respectively.

    Article Snippet: To detect gradual degradation of DNA fragments derived from the chromosome termini by treatment with Bal31 nuclease, total Chlorella chromosomal DNA (10 µg) or isolated minichromosomal DNA (1 µg) in 500 µl of 1× Bal31 buffer (20 mM Tris–HCl, pH 7.2, 0.6 M NaCl, 12.5 mM MgCl2 , 12.5 mM CaCl2 and 1 mM EDTA) was treated with 5 U Bal31 (Takara Shuzo) at 28°C for various time periods.

    Techniques: Derivative Assay