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Fluorescence intensity increment versus time of molecular beacon arrays immobilized on ( A ) a glutaraldehyde-derived glass slide and ( B ) an agarose film coated on a glass slide. The hybridization solution was 10 nM target T2 in 20 mM Tris–HCl pH 8.0 and 500 mM <t>MgCl2.</t>
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Article Title: Label-free hybridization detection of a single nucleotide mismatch by immobilization of molecular beacons on an agarose film

Journal: Nucleic Acids Research

doi:

Fluorescence intensity increment versus time of molecular beacon arrays immobilized on ( A ) a glutaraldehyde-derived glass slide and ( B ) an agarose film coated on a glass slide. The hybridization solution was 10 nM target T2 in 20 mM Tris–HCl pH 8.0 and 500 mM MgCl2.
Figure Legend Snippet: Fluorescence intensity increment versus time of molecular beacon arrays immobilized on ( A ) a glutaraldehyde-derived glass slide and ( B ) an agarose film coated on a glass slide. The hybridization solution was 10 nM target T2 in 20 mM Tris–HCl pH 8.0 and 500 mM MgCl2.

Techniques Used: Fluorescence, Derivative Assay, Hybridization

Hybridization dynamics images of molecular beacon arrays immobilized on ( A ) a glutaraldehyde-derived glass slide and ( B ) an agarose film. The hybridization solution was 10 nM target T2 in 20 mM Tris–HCl pH 8.0 and 500 mM MgCl2.
Figure Legend Snippet: Hybridization dynamics images of molecular beacon arrays immobilized on ( A ) a glutaraldehyde-derived glass slide and ( B ) an agarose film. The hybridization solution was 10 nM target T2 in 20 mM Tris–HCl pH 8.0 and 500 mM MgCl2.

Techniques Used: Hybridization, Derivative Assay

Mismatch discrimination ratios of the different molecular beacons in bulk solution, immobilized on a glutaraldehyde-derived glass slide and on an agarose film, respectively. Discrimination ratio is defined as (PM – MM)/PM. PM is the fluorescence increment of perfectly matched probes and MM is the average fluorescence increment of three 1 base mismatched probes. The hybridization solution was 10 nM target T2 in 20 mM Tris–HCl pH 8.0, with different concentrations of MgCl2.
Figure Legend Snippet: Mismatch discrimination ratios of the different molecular beacons in bulk solution, immobilized on a glutaraldehyde-derived glass slide and on an agarose film, respectively. Discrimination ratio is defined as (PM – MM)/PM. PM is the fluorescence increment of perfectly matched probes and MM is the average fluorescence increment of three 1 base mismatched probes. The hybridization solution was 10 nM target T2 in 20 mM Tris–HCl pH 8.0, with different concentrations of MgCl2.

Techniques Used: Derivative Assay, Fluorescence, Hybridization

Related Articles

Clone Assay:

Article Title: Definition of Eight Mulberry Species in the Genus Morus by Internal Transcribed Spacer-Based Phylogeny
Article Snippet: Polymerase chain reaction (PCR) amplification was conducted in a reaction mixture of 20 μL containing 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 1.25 mM or 2.5 mM MgCl2 (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China). .. The target fragment was gel-purified using the Takara Agarose Gel DNA Purification Kit (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China), cloned into the vector pUCm-T (Sangon, Shanghai, China) by AT cloning according to manufacturer’s instructions, and individual clones were sequenced by BGI-Shenzhen (Shenzhen, China).

Centrifugation:

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. Following staining, the cells were collected by centrifugation at 300 × g for 5 min at 4°C, resuspended in PBS and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, San Jose, CA, USA).

Amplification:

Article Title: Some dietary factors can modulate the effect of the zinc transporters 8 polymorphism on the risk of metabolic syndrome
Article Snippet: For the SNP mentioned, the PCR reaction was optimized at a 12.5 µl total volume containing 1 µl DNA template, 6.25 µl master mix containing MgCl2 , Smart Taq polymerase (CinnaGene Co, Iran), BSA 0.1% (TaKaRa, Japan), 1.8 µl total primers (containing 0.6 µl outers and 1.2 µl inners), 0.05 µl Mgcl2 , and 3.4 µl water. .. The PCR amplification was carried out with an initial denaturation at 94 °C for 6 minutes, denaturation at 95 °C for 30 seconds (35 cycles), 35 seconds of annealing at 64.8 °C (35 cycles), 55 seconds of extension at 72 °C, and an additional 10 minutes of extension at 72 °C at the end of the final cycle.

Article Title: Amplification of miniature inverted-repeat transposable elements and the associated impact on gene regulation and alternative splicing in mulberry (Morus notabilis)
Article Snippet: Each 20 μL reaction contained 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 2.5 mM MgCl2 (Takara Biotechnology Company, Dalian, China). .. The PCR amplification protocol was as follows: 94 °C for 4 min; 32 cycles of 94 °C for 30 s, 58 °C for 30 s, and 72 °C for 1 min; 72 °C for 7 min.

Article Title: Genetic diversity and structure of an endangered medicinal herb: implications for conservation
Article Snippet: Paragraph title: Microsatellite amplification ... PCRs were performed in a 15 μL volume containing 30–50 ng of template DNA, 0.5 μL dNTPs (20 mM), 1 μL 10× PCR buffer containing 25 mM MgCl2 (TAKARA, Japan), 0.25 μL forward and 1 μL reverse primers (8 pmol each), and 1 μL fluorescently labelled M13 primer (8 pmol; 6-FAM, VIC, PET and NED).

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: PCR amplification and HRM analysis were performed on Rotor Gene Q (Qiagen). .. PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µl SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 2 µL bisulfite converted DNA, and water.

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: Primer sequences for unmethylated DNA were as follows: forward primer 5′-TTTGTTGTTGTTGTAGTTTTGGG-3′, reverse primer 5′-TAAAATAAATAACACACATCTCTT-3′, with amplicon length 194 bp. .. MS-PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µL SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 1.5 µL bisulfite converted DNA, and water.

Article Title: Genetic diagnosis of a rare myrmecochorous species, Plagiorhegma dubium (Berberidaceae): Historical genetic bottlenecks and strong spatial structures among populations. Genetic diagnosis of a rare myrmecochorous species, Plagiorhegma dubium (Berberidaceae): Historical genetic bottlenecks and strong spatial structures among populations
Article Snippet: We performed PCRs in a volume of 25 μl containing 1 ul of 10 ng template DNA, 2 μl dNTPs (20 mM), 2.5 μl 10 × PCR buffer containing MgCl2 (Takara, Japan), fluorescently labeled (FAM, HEX, and NED) forward and reverse primers, 0.5 μl (10 pmol each). .. ABI 3730XL automated sequencer (Applied Biosystems) was used to separate out the amplified fragments.

Article Title: Label-free hybridization detection of a single nucleotide mismatch by immobilization of molecular beacons on an agarose film
Article Snippet: A 218 bp DNA fragment of the apoE gene was amplified with the primer sequences 5′-TCCAAGGAGCTGCAGGCGGCGCA (forward) and 5′-GCCCCGGCCTGGTACACTGCCA (reverse) as previously described ( ). .. The symmetric PCRs were performed in a total volume of 25 µl containing 75 mM Tris–HCl pH 9.0, 20 mM ammonium sulfate, 0.1 ml/l Tween, 1.5 mM MgCl2 , 500 nM each primer, 200 µM dNTPs, 100 ml/l DMSO and 1 U Taq polymerase (Takara Shuzo Co. Ltd).

Article Title: Definition of Eight Mulberry Species in the Genus Morus by Internal Transcribed Spacer-Based Phylogeny
Article Snippet: .. Polymerase chain reaction (PCR) amplification was conducted in a reaction mixture of 20 μL containing 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 1.25 mM or 2.5 mM MgCl2 (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China). .. The PCR amplification protocol was as follows: 94°C for 5 min, followed by 30 cycles of 94°C for 30 s, 58°C for 30 s, and 72°C for 1 min, followed by a final 10 min extension at 72°C.

Mass Spectrometry:

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: .. MS-PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µL SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 1.5 µL bisulfite converted DNA, and water. .. The cycling conditions were as follows: an initial denaturation step 95 °C for 5 min, 40 cycles of 95 °C for 20 s, 58 °C for 30 s and 70 °C for 35 s. Each run included a bisulfite-converted universal methylated and unmethylated DNA (Qiagen) and a no template control.

Cytometry:

Article Title: Analysis of the high-dose-range radioresistance of prostate cancer cells, including cancer stem cells, based on a stochastic model
Article Snippet: Flow cytometric analysis for detecting the CSCs To analyze the expression of the CSC markers, the cells were incubated in 100 μl phosphate-buffered saline without calcium chloride or magnesium chloride (PBS (–), TAKARA BIO INC.) containing 5% FBS and FITC anti-human CD44 (3 μl/106 cells) and PE anti-human CD133 (3 μl/106 cells) or respective mouse IgG1 isotype control antibodies (3 μl/106 cells) for 15 min at 4°C in the dark. .. After staining, the cells were centrifuged, resuspended in PBS (–), and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, Ltd, Tokyo, Japan).

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. Following staining, the cells were collected by centrifugation at 300 × g for 5 min at 4°C, resuspended in PBS and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, San Jose, CA, USA).

Blocking Assay:

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: .. To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. Following staining, the cells were collected by centrifugation at 300 × g for 5 min at 4°C, resuspended in PBS and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, San Jose, CA, USA).

Electrophoresis:

Article Title: Some dietary factors can modulate the effect of the zinc transporters 8 polymorphism on the risk of metabolic syndrome
Article Snippet: For the SNP mentioned, the PCR reaction was optimized at a 12.5 µl total volume containing 1 µl DNA template, 6.25 µl master mix containing MgCl2 , Smart Taq polymerase (CinnaGene Co, Iran), BSA 0.1% (TaKaRa, Japan), 1.8 µl total primers (containing 0.6 µl outers and 1.2 µl inners), 0.05 µl Mgcl2 , and 3.4 µl water. .. The PCR products were resolved by electrophoresis on a 1.8% agarose gel, a procedure that rendered three types of bands in heterozygotes (391, 334, and 104 bp) and two types of bands in homozygotes (mutant allele carrier resulting in 391 and 104 bp, wild carrier allele resulting in 391 and 334 bp).

Article Title: An increasing trend of neonatal invasive multidrug-resistant group B streptococcus infections in southern China, 2011–2017
Article Snippet: The PCR was conducted in a final volume of 50 µL of 0.8× PCR buffer (50 mM KCl, 10 mM Tris-HCl [pH 9.0], 0.1% Triton X-100, 0.01% [wt/vol] stabilizer, 1.5 mM MgCl2 ) containing 300 µM deoxynucleoside triphosphates, 3.5 mM MgCl2 , 2 U Taq polymerase (Takara, Kusatsu, Japan), and 10 ng of template DNA, extracted by Lysis Buffer for Microorganism for direct PCR (Takara); primer concentrations were all 0.5 µM. .. The PCR products were analyzed by electrophoresis on a 2% agarose gel at 220 V for 45 minutes in 0.5× Tris-borate-EDTA buffer., Visualization and image acquisition were performed with gold view (SBS, Beijing, China) and ultra-violet transillumination.

Incubation:

Article Title: Analysis of the high-dose-range radioresistance of prostate cancer cells, including cancer stem cells, based on a stochastic model
Article Snippet: .. Flow cytometric analysis for detecting the CSCs To analyze the expression of the CSC markers, the cells were incubated in 100 μl phosphate-buffered saline without calcium chloride or magnesium chloride (PBS (–), TAKARA BIO INC.) containing 5% FBS and FITC anti-human CD44 (3 μl/106 cells) and PE anti-human CD133 (3 μl/106 cells) or respective mouse IgG1 isotype control antibodies (3 μl/106 cells) for 15 min at 4°C in the dark. .. After staining, the cells were centrifuged, resuspended in PBS (–), and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, Ltd, Tokyo, Japan).

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: .. To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. Following staining, the cells were collected by centrifugation at 300 × g for 5 min at 4°C, resuspended in PBS and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, San Jose, CA, USA).

Formalin-fixed Paraffin-Embedded:

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: Primers were designed using MethPrimer, considering the fact that FFPE DNA is highly fragmented and amplicons over 200 bp in length result in lower melting resolution. .. PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µl SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 2 µL bisulfite converted DNA, and water.

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: Primers were designed using MethPrimer again with consideration of the FFPE DNA fragmentation. .. MS-PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µL SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 1.5 µL bisulfite converted DNA, and water.

Expressing:

Article Title: Analysis of the high-dose-range radioresistance of prostate cancer cells, including cancer stem cells, based on a stochastic model
Article Snippet: .. Flow cytometric analysis for detecting the CSCs To analyze the expression of the CSC markers, the cells were incubated in 100 μl phosphate-buffered saline without calcium chloride or magnesium chloride (PBS (–), TAKARA BIO INC.) containing 5% FBS and FITC anti-human CD44 (3 μl/106 cells) and PE anti-human CD133 (3 μl/106 cells) or respective mouse IgG1 isotype control antibodies (3 μl/106 cells) for 15 min at 4°C in the dark. .. After staining, the cells were centrifuged, resuspended in PBS (–), and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, Ltd, Tokyo, Japan).

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: .. To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. Following staining, the cells were collected by centrifugation at 300 × g for 5 min at 4°C, resuspended in PBS and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, San Jose, CA, USA).

Hybridization:

Article Title: Label-free hybridization detection of a single nucleotide mismatch by immobilization of molecular beacons on an agarose film
Article Snippet: Paragraph title: Hybridization of PCR products to the molecular beacon array ... The symmetric PCRs were performed in a total volume of 25 µl containing 75 mM Tris–HCl pH 9.0, 20 mM ammonium sulfate, 0.1 ml/l Tween, 1.5 mM MgCl2 , 500 nM each primer, 200 µM dNTPs, 100 ml/l DMSO and 1 U Taq polymerase (Takara Shuzo Co. Ltd).

Countercurrent Chromatography:

Article Title: Some dietary factors can modulate the effect of the zinc transporters 8 polymorphism on the risk of metabolic syndrome
Article Snippet: Our T-ARMS assay with different inner allele specific primers produced allele-specific PCR products; forward: CTT CTT TAT CAA CAG CAG CCC GCC, and reverse: TCT CCG AAC CAC TAG GCT GTA CCA. .. For the SNP mentioned, the PCR reaction was optimized at a 12.5 µl total volume containing 1 µl DNA template, 6.25 µl master mix containing MgCl2 , Smart Taq polymerase (CinnaGene Co, Iran), BSA 0.1% (TaKaRa, Japan), 1.8 µl total primers (containing 0.6 µl outers and 1.2 µl inners), 0.05 µl Mgcl2 , and 3.4 µl water.

Flow Cytometry:

Article Title: Analysis of the high-dose-range radioresistance of prostate cancer cells, including cancer stem cells, based on a stochastic model
Article Snippet: .. Flow cytometric analysis for detecting the CSCs To analyze the expression of the CSC markers, the cells were incubated in 100 μl phosphate-buffered saline without calcium chloride or magnesium chloride (PBS (–), TAKARA BIO INC.) containing 5% FBS and FITC anti-human CD44 (3 μl/106 cells) and PE anti-human CD133 (3 μl/106 cells) or respective mouse IgG1 isotype control antibodies (3 μl/106 cells) for 15 min at 4°C in the dark. .. After staining, the cells were centrifuged, resuspended in PBS (–), and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, Ltd, Tokyo, Japan).

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: Paragraph title: Flow cytometric analysis ... To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark.

Sequencing:

Article Title: Interaction between TP53 and XRCC1 increases susceptibility to cervical cancer development: a case control study
Article Snippet: The rs121913483 (C > G), rs25487 (G > A), and rs1042522 (G > C) genotypes were identified separately by MAMA-PCR and then validated by Sanger sequencing (Sangon Biotech, Shanghai, China) as described in Additional file : Figure S1. .. The PCR primers (Invitrogen, Shanghai, China) used to amplify the genes are as follows: The final reaction mixture contained 50 ng template DNA, 0.2 μL Taq polymerase, 2.4 μL dNTPs, 2 μL 10X PCR Buffer (Takara, Japan), 50 nmol each reverse and forward primers, MgCl2 (Takara, Japan; FGFR3, 1 μL; XRCC1, 1.5 μL; TP53, 2 μL), and double-distilled H2 O at a final reaction volume of 20 μL.

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: Sequence of forward primer was 5′-TTTTGGATTAAAGYGGAATTGAG-3′, sequence of reverse primer 5′-TCCATTATACTCACRCTAAAAAAC-3′, with amplicon length 153 bp. .. PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µl SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 2 µL bisulfite converted DNA, and water.

Article Title: Insulin resistance in cavefish as an adaptation to a nutrient-limited environment
Article Snippet: PCR reactions were carried out in 12.5-µl volume containing 1× LA PCR Buffer II (Clontech), 2.5 mM MgCl2 , 0.4 mM dNTP mix, 0.4 µM of each forward and reverse primer and 0.05 units of TaKaRa LA Taq DNA Polymerase (Clontech). .. The PCR products were diluted tenfold and sequenced directly on a 3730XL DNA Analyzer (Applied Biosystems) using the sequencing primer: GGTGGAGTTGATGGTGGTATAG.

Article Title: Definition of Eight Mulberry Species in the Genus Morus by Internal Transcribed Spacer-Based Phylogeny
Article Snippet: Paragraph title: DNA isolation, amplification, and sequencing ... Polymerase chain reaction (PCR) amplification was conducted in a reaction mixture of 20 μL containing 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 1.25 mM or 2.5 mM MgCl2 (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China).

Cellular Antioxidant Activity Assay:

Article Title: Some dietary factors can modulate the effect of the zinc transporters 8 polymorphism on the risk of metabolic syndrome
Article Snippet: Our T-ARMS assay with different inner allele specific primers produced allele-specific PCR products; forward: CTT CTT TAT CAA CAG CAG CCC GCC, and reverse: TCT CCG AAC CAC TAG GCT GTA CCA. .. For the SNP mentioned, the PCR reaction was optimized at a 12.5 µl total volume containing 1 µl DNA template, 6.25 µl master mix containing MgCl2 , Smart Taq polymerase (CinnaGene Co, Iran), BSA 0.1% (TaKaRa, Japan), 1.8 µl total primers (containing 0.6 µl outers and 1.2 µl inners), 0.05 µl Mgcl2 , and 3.4 µl water.

Immunofluorescence:

Article Title: Analysis of the high-dose-range radioresistance of prostate cancer cells, including cancer stem cells, based on a stochastic model
Article Snippet: Flow cytometric analysis for detecting the CSCs To analyze the expression of the CSC markers, the cells were incubated in 100 μl phosphate-buffered saline without calcium chloride or magnesium chloride (PBS (–), TAKARA BIO INC.) containing 5% FBS and FITC anti-human CD44 (3 μl/106 cells) and PE anti-human CD133 (3 μl/106 cells) or respective mouse IgG1 isotype control antibodies (3 μl/106 cells) for 15 min at 4°C in the dark. .. After staining, the cells were centrifuged, resuspended in PBS (–), and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, Ltd, Tokyo, Japan).

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. Following staining, the cells were collected by centrifugation at 300 × g for 5 min at 4°C, resuspended in PBS and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, San Jose, CA, USA).

DNA Extraction:

Article Title: Interaction between TP53 and XRCC1 increases susceptibility to cervical cancer development: a case control study
Article Snippet: Genomic DNA was extracted from peripheral blood samples using a Rapid Blood Genomic DNA Isolation Kit (Sangon Biotech, Shanghai, China), and extracts were stored at − 20 °C until use. .. The PCR primers (Invitrogen, Shanghai, China) used to amplify the genes are as follows: The final reaction mixture contained 50 ng template DNA, 0.2 μL Taq polymerase, 2.4 μL dNTPs, 2 μL 10X PCR Buffer (Takara, Japan), 50 nmol each reverse and forward primers, MgCl2 (Takara, Japan; FGFR3, 1 μL; XRCC1, 1.5 μL; TP53, 2 μL), and double-distilled H2 O at a final reaction volume of 20 μL.

Article Title: Definition of Eight Mulberry Species in the Genus Morus by Internal Transcribed Spacer-Based Phylogeny
Article Snippet: Paragraph title: DNA isolation, amplification, and sequencing ... Polymerase chain reaction (PCR) amplification was conducted in a reaction mixture of 20 μL containing 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 1.25 mM or 2.5 mM MgCl2 (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China).

Fluorescence:

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. The fluorescence values of the respective isotype controls were subtracted from the fluorescence data of CD44, CD133 and CD138.

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: MS-PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µL SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 1.5 µL bisulfite converted DNA, and water. .. Fluorescence data were analyzed using Rotor gene Q software.

Methylation:

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µl SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 2 µL bisulfite converted DNA, and water. .. Each run included a no template control, a bisulfite-converted universal methylated and unmethylated DNA (Qiagen) and prepared standard containing 10% of universal methylated DNA which served as a cut-off for methylation status.

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: Paragraph title: 4.5. Real-Time Methylation Specific Analysis of GATA4 Gene ... MS-PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µL SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 1.5 µL bisulfite converted DNA, and water.

Mutagenesis:

Article Title: Some dietary factors can modulate the effect of the zinc transporters 8 polymorphism on the risk of metabolic syndrome
Article Snippet: The selected polymorphism (rs13266634) was studied using the tetra-primer refractory mutation system-polymerase chain reaction (T-ARMS-PCR) method. .. For the SNP mentioned, the PCR reaction was optimized at a 12.5 µl total volume containing 1 µl DNA template, 6.25 µl master mix containing MgCl2 , Smart Taq polymerase (CinnaGene Co, Iran), BSA 0.1% (TaKaRa, Japan), 1.8 µl total primers (containing 0.6 µl outers and 1.2 µl inners), 0.05 µl Mgcl2 , and 3.4 µl water.

Isolation:

Article Title: Interaction between TP53 and XRCC1 increases susceptibility to cervical cancer development: a case control study
Article Snippet: Paragraph title: Isolation of genomic DNA ... The PCR primers (Invitrogen, Shanghai, China) used to amplify the genes are as follows: The final reaction mixture contained 50 ng template DNA, 0.2 μL Taq polymerase, 2.4 μL dNTPs, 2 μL 10X PCR Buffer (Takara, Japan), 50 nmol each reverse and forward primers, MgCl2 (Takara, Japan; FGFR3, 1 μL; XRCC1, 1.5 μL; TP53, 2 μL), and double-distilled H2 O at a final reaction volume of 20 μL.

Labeling:

Article Title: Genetic diagnosis of a rare myrmecochorous species, Plagiorhegma dubium (Berberidaceae): Historical genetic bottlenecks and strong spatial structures among populations. Genetic diagnosis of a rare myrmecochorous species, Plagiorhegma dubium (Berberidaceae): Historical genetic bottlenecks and strong spatial structures among populations
Article Snippet: .. We performed PCRs in a volume of 25 μl containing 1 ul of 10 ng template DNA, 2 μl dNTPs (20 mM), 2.5 μl 10 × PCR buffer containing MgCl2 (Takara, Japan), fluorescently labeled (FAM, HEX, and NED) forward and reverse primers, 0.5 μl (10 pmol each). .. PCR cycling conditions were as follows: 5 min predenaturation at 95°C followed by 30 cycles of 1 min at 95°C, 1 min at 50°C, and 1 min at 72°C, followed by a final 10 min extension step at 72°C.

Polymerase Chain Reaction:

Article Title: Some dietary factors can modulate the effect of the zinc transporters 8 polymorphism on the risk of metabolic syndrome
Article Snippet: .. For the SNP mentioned, the PCR reaction was optimized at a 12.5 µl total volume containing 1 µl DNA template, 6.25 µl master mix containing MgCl2 , Smart Taq polymerase (CinnaGene Co, Iran), BSA 0.1% (TaKaRa, Japan), 1.8 µl total primers (containing 0.6 µl outers and 1.2 µl inners), 0.05 µl Mgcl2 , and 3.4 µl water. .. The PCR amplification was carried out with an initial denaturation at 94 °C for 6 minutes, denaturation at 95 °C for 30 seconds (35 cycles), 35 seconds of annealing at 64.8 °C (35 cycles), 55 seconds of extension at 72 °C, and an additional 10 minutes of extension at 72 °C at the end of the final cycle.

Article Title: Amplification of miniature inverted-repeat transposable elements and the associated impact on gene regulation and alternative splicing in mulberry (Morus notabilis)
Article Snippet: .. Each 20 μL reaction contained 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 2.5 mM MgCl2 (Takara Biotechnology Company, Dalian, China). .. The PCR amplification protocol was as follows: 94 °C for 4 min; 32 cycles of 94 °C for 30 s, 58 °C for 30 s, and 72 °C for 1 min; 72 °C for 7 min.

Article Title: An increasing trend of neonatal invasive multidrug-resistant group B streptococcus infections in southern China, 2011–2017
Article Snippet: .. The PCR was conducted in a final volume of 50 µL of 0.8× PCR buffer (50 mM KCl, 10 mM Tris-HCl [pH 9.0], 0.1% Triton X-100, 0.01% [wt/vol] stabilizer, 1.5 mM MgCl2 ) containing 300 µM deoxynucleoside triphosphates, 3.5 mM MgCl2 , 2 U Taq polymerase (Takara, Kusatsu, Japan), and 10 ng of template DNA, extracted by Lysis Buffer for Microorganism for direct PCR (Takara); primer concentrations were all 0.5 µM. .. The PCR was conducted on a DNA thermal cycler (Bio-Rad, Hercules, CA, USA) with the following cycling conditions: an initial cycle of 3 minutes at 94°C (5 minutes for tetO ); 30 cycles of 30 seconds of denaturation at 94°C, 30 seconds of annealing at 55°C, and 50 seconds of extension at 72°C (45 seconds for tetO ); followed by 33 cycles of elongation at 72°C.

Article Title: Genetic diversity and structure of an endangered medicinal herb: implications for conservation
Article Snippet: .. PCRs were performed in a 15 μL volume containing 30–50 ng of template DNA, 0.5 μL dNTPs (20 mM), 1 μL 10× PCR buffer containing 25 mM MgCl2 (TAKARA, Japan), 0.25 μL forward and 1 μL reverse primers (8 pmol each), and 1 μL fluorescently labelled M13 primer (8 pmol; 6-FAM, VIC, PET and NED). .. PCR cycling conditions were as follows: 5 min pre-denaturation at 94 °C followed by 30 cycles of 30 s at 94 °C, 45 s at 56 °C and 45 s at 72 °C, followed by eight cycles of 30 s at 94 °C, 45 s at 53 °C and 45 s at 72 °C, and then a final 20 min extension step at 72 °C.

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: .. PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µl SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 2 µL bisulfite converted DNA, and water. ..

Article Title: Genetic diagnosis of a rare myrmecochorous species, Plagiorhegma dubium (Berberidaceae): Historical genetic bottlenecks and strong spatial structures among populations. Genetic diagnosis of a rare myrmecochorous species, Plagiorhegma dubium (Berberidaceae): Historical genetic bottlenecks and strong spatial structures among populations
Article Snippet: .. We performed PCRs in a volume of 25 μl containing 1 ul of 10 ng template DNA, 2 μl dNTPs (20 mM), 2.5 μl 10 × PCR buffer containing MgCl2 (Takara, Japan), fluorescently labeled (FAM, HEX, and NED) forward and reverse primers, 0.5 μl (10 pmol each). .. PCR cycling conditions were as follows: 5 min predenaturation at 95°C followed by 30 cycles of 1 min at 95°C, 1 min at 50°C, and 1 min at 72°C, followed by a final 10 min extension step at 72°C.

Article Title: Insulin resistance in cavefish as an adaptation to a nutrient-limited environment
Article Snippet: .. PCR reactions were carried out in 12.5-µl volume containing 1× LA PCR Buffer II (Clontech), 2.5 mM MgCl2 , 0.4 mM dNTP mix, 0.4 µM of each forward and reverse primer and 0.05 units of TaKaRa LA Taq DNA Polymerase (Clontech). .. The PCR cycling conditions were as follows: initial denaturation at 94 °C for 2 min, followed by 35 cycles of 94 °C for 30 s, annealing temperature 52 °C for 30 s and 72 °C for 1 min. A final 5-min elongation step was performed at 72 °C.

Article Title: Label-free hybridization detection of a single nucleotide mismatch by immobilization of molecular beacons on an agarose film
Article Snippet: Paragraph title: Hybridization of PCR products to the molecular beacon array ... The symmetric PCRs were performed in a total volume of 25 µl containing 75 mM Tris–HCl pH 9.0, 20 mM ammonium sulfate, 0.1 ml/l Tween, 1.5 mM MgCl2 , 500 nM each primer, 200 µM dNTPs, 100 ml/l DMSO and 1 U Taq polymerase (Takara Shuzo Co. Ltd).

Article Title: Definition of Eight Mulberry Species in the Genus Morus by Internal Transcribed Spacer-Based Phylogeny
Article Snippet: .. Polymerase chain reaction (PCR) amplification was conducted in a reaction mixture of 20 μL containing 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 1.25 mM or 2.5 mM MgCl2 (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China). .. The PCR amplification protocol was as follows: 94°C for 5 min, followed by 30 cycles of 94°C for 30 s, 58°C for 30 s, and 72°C for 1 min, followed by a final 10 min extension at 72°C.

FACS:

Article Title: Analysis of the high-dose-range radioresistance of prostate cancer cells, including cancer stem cells, based on a stochastic model
Article Snippet: Flow cytometric analysis for detecting the CSCs To analyze the expression of the CSC markers, the cells were incubated in 100 μl phosphate-buffered saline without calcium chloride or magnesium chloride (PBS (–), TAKARA BIO INC.) containing 5% FBS and FITC anti-human CD44 (3 μl/106 cells) and PE anti-human CD133 (3 μl/106 cells) or respective mouse IgG1 isotype control antibodies (3 μl/106 cells) for 15 min at 4°C in the dark. .. After staining, the cells were centrifuged, resuspended in PBS (–), and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, Ltd, Tokyo, Japan).

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. Following staining, the cells were collected by centrifugation at 300 × g for 5 min at 4°C, resuspended in PBS and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, San Jose, CA, USA).

Salting Out:

Article Title: Some dietary factors can modulate the effect of the zinc transporters 8 polymorphism on the risk of metabolic syndrome
Article Snippet: Hence, the genomic DNA was extracted from the peripheral blood using standard salting-out method . .. For the SNP mentioned, the PCR reaction was optimized at a 12.5 µl total volume containing 1 µl DNA template, 6.25 µl master mix containing MgCl2 , Smart Taq polymerase (CinnaGene Co, Iran), BSA 0.1% (TaKaRa, Japan), 1.8 µl total primers (containing 0.6 µl outers and 1.2 µl inners), 0.05 µl Mgcl2 , and 3.4 µl water.

Activated Clotting Time Assay:

Article Title: Definition of Eight Mulberry Species in the Genus Morus by Internal Transcribed Spacer-Based Phylogeny
Article Snippet: The target sequence regions (the partial 18S rRNA gene, ITS1, 5.8S rRNA gene, ITS2, and the partial 28S rRNA gene) were amplified with mulberry-specific forward ( 5′-GTA ACA AGG TTT CCG TAG GTG-3′ ) and reverse ( 5′-TAA ACT CAG CGG GTA GCC-3′ ) primers, which were designed from the highly conserved regions flanking the 3′ end of the 18S rRNA gene and the 5′ end of the 28S rRNA gene, respectively, based on available Morus rRNA gene sequences in GenBank (National Center for Biotechnology Information (NCBI), http://www.ncbi.nlm.nih.gov/ ). .. Polymerase chain reaction (PCR) amplification was conducted in a reaction mixture of 20 μL containing 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 1.25 mM or 2.5 mM MgCl2 (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China).

Plasmid Preparation:

Article Title: Definition of Eight Mulberry Species in the Genus Morus by Internal Transcribed Spacer-Based Phylogeny
Article Snippet: Polymerase chain reaction (PCR) amplification was conducted in a reaction mixture of 20 μL containing 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 1.25 mM or 2.5 mM MgCl2 (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China). .. The target fragment was gel-purified using the Takara Agarose Gel DNA Purification Kit (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China), cloned into the vector pUCm-T (Sangon, Shanghai, China) by AT cloning according to manufacturer’s instructions, and individual clones were sequenced by BGI-Shenzhen (Shenzhen, China).

Software:

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. The results were analyzed using the Kaluza software (version 1.5a; Beckman Coulter, Inc., Brea, CA, USA).

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µl SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 2 µL bisulfite converted DNA, and water. .. HRM data were analyzed using Rotor gene Q software 2.3 (Qiagen).

Article Title: Next-Generation Sequencing Approach in Methylation Analysis of HNF1B and GATA4 Genes: Searching for Biomarkers in Ovarian Cancer
Article Snippet: MS-PCR was carried out in a final volume of 20 µL containing: 2 µL 10× Reaction Buffer no MgCl2 , 2 µL MgCl2 (25 mM), 1.6 µL dNTPs solution Takara (2.5 mM), 0.5 µL of each primer (10 µM), 0.3 µL SYTO® 9 Dye (0.05 mM), 0.25 µL AmpliTaq Gold® DNA Polymerase (Thermo Fisher Scientific), 1.5 µL bisulfite converted DNA, and water. .. Fluorescence data were analyzed using Rotor gene Q software.

Positron Emission Tomography:

Article Title: Genetic diversity and structure of an endangered medicinal herb: implications for conservation
Article Snippet: .. PCRs were performed in a 15 μL volume containing 30–50 ng of template DNA, 0.5 μL dNTPs (20 mM), 1 μL 10× PCR buffer containing 25 mM MgCl2 (TAKARA, Japan), 0.25 μL forward and 1 μL reverse primers (8 pmol each), and 1 μL fluorescently labelled M13 primer (8 pmol; 6-FAM, VIC, PET and NED). .. PCR cycling conditions were as follows: 5 min pre-denaturation at 94 °C followed by 30 cycles of 30 s at 94 °C, 45 s at 56 °C and 45 s at 72 °C, followed by eight cycles of 30 s at 94 °C, 45 s at 53 °C and 45 s at 72 °C, and then a final 20 min extension step at 72 °C.

Agarose Gel Electrophoresis:

Article Title: Some dietary factors can modulate the effect of the zinc transporters 8 polymorphism on the risk of metabolic syndrome
Article Snippet: For the SNP mentioned, the PCR reaction was optimized at a 12.5 µl total volume containing 1 µl DNA template, 6.25 µl master mix containing MgCl2 , Smart Taq polymerase (CinnaGene Co, Iran), BSA 0.1% (TaKaRa, Japan), 1.8 µl total primers (containing 0.6 µl outers and 1.2 µl inners), 0.05 µl Mgcl2 , and 3.4 µl water. .. The PCR products were resolved by electrophoresis on a 1.8% agarose gel, a procedure that rendered three types of bands in heterozygotes (391, 334, and 104 bp) and two types of bands in homozygotes (mutant allele carrier resulting in 391 and 104 bp, wild carrier allele resulting in 391 and 334 bp).

Article Title: An increasing trend of neonatal invasive multidrug-resistant group B streptococcus infections in southern China, 2011–2017
Article Snippet: The PCR was conducted in a final volume of 50 µL of 0.8× PCR buffer (50 mM KCl, 10 mM Tris-HCl [pH 9.0], 0.1% Triton X-100, 0.01% [wt/vol] stabilizer, 1.5 mM MgCl2 ) containing 300 µM deoxynucleoside triphosphates, 3.5 mM MgCl2 , 2 U Taq polymerase (Takara, Kusatsu, Japan), and 10 ng of template DNA, extracted by Lysis Buffer for Microorganism for direct PCR (Takara); primer concentrations were all 0.5 µM. .. The PCR products were analyzed by electrophoresis on a 2% agarose gel at 220 V for 45 minutes in 0.5× Tris-borate-EDTA buffer., Visualization and image acquisition were performed with gold view (SBS, Beijing, China) and ultra-violet transillumination.

Article Title: Interaction between TP53 and XRCC1 increases susceptibility to cervical cancer development: a case control study
Article Snippet: DNA quantity was measured using an ultraviolet spectrophotometer at 260 nm, and the DNA quality was assessed by agarose gel electrophoresis. .. The PCR primers (Invitrogen, Shanghai, China) used to amplify the genes are as follows: The final reaction mixture contained 50 ng template DNA, 0.2 μL Taq polymerase, 2.4 μL dNTPs, 2 μL 10X PCR Buffer (Takara, Japan), 50 nmol each reverse and forward primers, MgCl2 (Takara, Japan; FGFR3, 1 μL; XRCC1, 1.5 μL; TP53, 2 μL), and double-distilled H2 O at a final reaction volume of 20 μL.

Spectrophotometry:

Article Title: Interaction between TP53 and XRCC1 increases susceptibility to cervical cancer development: a case control study
Article Snippet: DNA quantity was measured using an ultraviolet spectrophotometer at 260 nm, and the DNA quality was assessed by agarose gel electrophoresis. .. The PCR primers (Invitrogen, Shanghai, China) used to amplify the genes are as follows: The final reaction mixture contained 50 ng template DNA, 0.2 μL Taq polymerase, 2.4 μL dNTPs, 2 μL 10X PCR Buffer (Takara, Japan), 50 nmol each reverse and forward primers, MgCl2 (Takara, Japan; FGFR3, 1 μL; XRCC1, 1.5 μL; TP53, 2 μL), and double-distilled H2 O at a final reaction volume of 20 μL.

Produced:

Article Title: Some dietary factors can modulate the effect of the zinc transporters 8 polymorphism on the risk of metabolic syndrome
Article Snippet: Two outer primers produced a PCR product, which is to be used as an internal control for reaction; forward: GAA GTT GGA GTC AGA GCA GTC GCC, and reverse: ATC TCA GTG CCT CTT CCT TCA TGG TGA. .. For the SNP mentioned, the PCR reaction was optimized at a 12.5 µl total volume containing 1 µl DNA template, 6.25 µl master mix containing MgCl2 , Smart Taq polymerase (CinnaGene Co, Iran), BSA 0.1% (TaKaRa, Japan), 1.8 µl total primers (containing 0.6 µl outers and 1.2 µl inners), 0.05 µl Mgcl2 , and 3.4 µl water.

Concentration Assay:

Article Title: Label-free hybridization detection of a single nucleotide mismatch by immobilization of molecular beacons on an agarose film
Article Snippet: The symmetric PCRs were performed in a total volume of 25 µl containing 75 mM Tris–HCl pH 9.0, 20 mM ammonium sulfate, 0.1 ml/l Tween, 1.5 mM MgCl2 , 500 nM each primer, 200 µM dNTPs, 100 ml/l DMSO and 1 U Taq polymerase (Takara Shuzo Co. Ltd). .. The mixture for asymmetric PCR contained all the reaction components in identical amounts as in the symmetric PCR except that the forward primer concentration was 50 nM.

DNA Purification:

Article Title: Definition of Eight Mulberry Species in the Genus Morus by Internal Transcribed Spacer-Based Phylogeny
Article Snippet: Polymerase chain reaction (PCR) amplification was conducted in a reaction mixture of 20 μL containing 20 ng genomic DNA, 2 mM PCR buffer, 0.2 mM each primer, 0.2 mM each dNTP, and 1 unit Taq polymerase with 1.25 mM or 2.5 mM MgCl2 (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China). .. The target fragment was gel-purified using the Takara Agarose Gel DNA Purification Kit (Takara Bio-technology (Dalian) Co. Ltd., Dalian, China), cloned into the vector pUCm-T (Sangon, Shanghai, China) by AT cloning according to manufacturer’s instructions, and individual clones were sequenced by BGI-Shenzhen (Shenzhen, China).

Lysis:

Article Title: An increasing trend of neonatal invasive multidrug-resistant group B streptococcus infections in southern China, 2011–2017
Article Snippet: .. The PCR was conducted in a final volume of 50 µL of 0.8× PCR buffer (50 mM KCl, 10 mM Tris-HCl [pH 9.0], 0.1% Triton X-100, 0.01% [wt/vol] stabilizer, 1.5 mM MgCl2 ) containing 300 µM deoxynucleoside triphosphates, 3.5 mM MgCl2 , 2 U Taq polymerase (Takara, Kusatsu, Japan), and 10 ng of template DNA, extracted by Lysis Buffer for Microorganism for direct PCR (Takara); primer concentrations were all 0.5 µM. .. The PCR was conducted on a DNA thermal cycler (Bio-Rad, Hercules, CA, USA) with the following cycling conditions: an initial cycle of 3 minutes at 94°C (5 minutes for tetO ); 30 cycles of 30 seconds of denaturation at 94°C, 30 seconds of annealing at 55°C, and 50 seconds of extension at 72°C (45 seconds for tetO ); followed by 33 cycles of elongation at 72°C.

Staining:

Article Title: Analysis of the high-dose-range radioresistance of prostate cancer cells, including cancer stem cells, based on a stochastic model
Article Snippet: Flow cytometric analysis for detecting the CSCs To analyze the expression of the CSC markers, the cells were incubated in 100 μl phosphate-buffered saline without calcium chloride or magnesium chloride (PBS (–), TAKARA BIO INC.) containing 5% FBS and FITC anti-human CD44 (3 μl/106 cells) and PE anti-human CD133 (3 μl/106 cells) or respective mouse IgG1 isotype control antibodies (3 μl/106 cells) for 15 min at 4°C in the dark. .. After staining, the cells were centrifuged, resuspended in PBS (–), and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, Ltd, Tokyo, Japan).

Article Title: Understanding the mechanism underlying the acquisition of radioresistance in human prostate cancer cells
Article Snippet: To analyze the expression of the CSC markers, the cells were incubated in 100 µl PBS without calcium chloride and magnesium chloride [PBS(−); Takara Bio Inc.] containing 5% FBS used as blocking agent and PE anti-human CD44 (3 µl/106 cells), CD133/1-APC (3 µl:106 cells), PerCP/Cy5.5 anti-human CD138 (3 µl/106 cells) or respective mouse IgG1 isotype control antibodies (3 µl/106 cells) for 15 min at 4°C in the dark. .. Following staining, the cells were collected by centrifugation at 300 × g for 5 min at 4°C, resuspended in PBS and analyzed by direct immunofluorescence flow cytometry using a BD FACS Aria™ Cell Sorter (BD Biosciences, San Jose, CA, USA).

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