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Boehringer Mannheim dntp
Dntp, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 92/100, based on 95 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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dntp - by Bioz Stars, 2020-04
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Clone Assay:

Article Title: Major Protein of Resting Rhizomes of Calystegia sepium (Hedge Bindweed) Closely Resembles Plant RNases But Has No Enzymatic Activity 1
Article Snippet: The reaction mixture for amplification of genomic DNA sequences contained 10 m m Tris-HCl, pH 8.3, 50 m m KCl, 1.5 m m MgCl2 , 100 mg/L gelatin, 0.4 m m of each dNTP, 2.5 units of Taq polymerase (Boehringer Mannheim, Basel), 50 ng to 500 μg of genomic DNA, and 20 μL of the appropriate primer mixture (20 μ m ) in a 100-μL reaction volume. .. The PCR primers were derived from both ends of the coding sequence of the cDNA clones encoding CalsepRRP.

Amplification:

Article Title: Induction of neuropilin-1 and vascular endothelial growth factor by epidermal growth factor in human gastric cancer cells
Article Snippet: For reverse transcriptase–polymerase chain reaction (RT–PCR), 3 μ g of total RNA was used for cDNA synthesis with avian myeloblastosis virus reverse transcriptase (Life Technologies) in a final volume of 20 μ l. The reaction mixture included 0.5 M Tris-HCl (pH 8.0), 0.5 M KCl, 0.05 M MgCl2 , 2.5 mM dNTP, 40 U of RNase inhibitor (Boehringer Mannheim, Indianapolis, IN, USA), 50 U of reverse transcriptase, and 0.5 μ g of random primers. .. A portion of the reaction mixture (5 μ l) was subjected to PCR amplification in a reaction mixture (50 μ l) that contained 1 μ mol l−1 of each of two primers (sense and antisense), 1.5 mmol l−1 of MgCl2 , 0.2 mmol l−1 of each of four deoxynucleotides, and 2.5 U of Taq polymerase (Promega, Madison, WI, USA).

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.) .. PCR cycling consisted of initial denaturation at 95°C for 30 s, followed by 35 cycles consisting of denaturation at 95°C for 30 s, primer annealing at 50°C ( / GC ) or 60°C ( GC / ) for 30 s, and extension at 72°C for 30 s. Prior to DGGE analysis, the PCR products were heated to 95°C to denature the DNA and then were slowly cooled to room temperature to allow for reannealing of the DNA strands.

Article Title: Serum levels of the angiogenic factor pleiotrophin in relation to disease stage in lung cancer patients
Article Snippet: .. PCR The cDNA (1.5 μl) was amplified in a 30 μl final volume of 10 mM Tris (pH 8.3), 50 mM KCl, 1.5 mM MgCl2 , 200 μl of each dNTP, 0.5 μM of each primer and 1.25 U Taq polymerase (Boehringer Mannheim, Germany) for 25–30 cycles under the following conditions: 5 min at 95°C, 1 min at 56.5°C (PTN) or at 58°C (GAPDH) and 2 min at 73°C. ..

Article Title: Polymorphisms of Human Leukocyte Antigen Genes in Korean Children with Kawasaki Disease
Article Snippet: HLA-A, -B, and -C Genotyping The genotyping of HLA-A, -B, and -C was performed by the amplification refractory mutation system (ARMS)–polymerase chain reaction (PCR) method. .. The PCR procedure was carried out in a reaction (13 μl), containing 100 to 200 ng genomic DNA, 0.8 X buffer (40 mmol/l KCl, 1.2 mmol/l MgCl2 , 8 mmol/l Tris-HCl pH 8.8, 0.08% Triton X-100), 5% dimethylsulphoxide (DMSO), 200 μmol/l of each dNTP, 0.25 U Taq DNA polymerase (Boehringer, Mannheim, Germany), 1 μmol/l of each sequence-specific primer, and 0.2 μmol/l of internal control primers.

Article Title: Immunological and pathological comparative analysis between experimental latent tuberculous infection and progressive pulmonary tuberculosis
Article Snippet: The PCR was carried out by incubating the sections with 50 μl of 1× reaction buffer (G ibco BRL), 1·5 U Taq polymerase, 2 m m MgCl2 , 40 μ m dNTP, 0·2 μ m dUTP labelled with digoxigenin (Boehringer Manheim Indianapolis, IN, USA) and 60 pg each of IS6110 m tuberculosis insertion sequence primers. .. The M. tuberculosis DNA amplification started with denaturation at 94°C for 1 min, annealing at 70°C for 1 min and extension at 72°C for 1 min, for 35 cycles.

Article Title: Characterization of SRp46, a Novel Human SR Splicing Factor Encoded by a PR264/SC35 Retropseudogene
Article Snippet: .. Double-stranded cDNA was amplified in the presence of a 0.8 μM concentration of each amplimer (H430-1507 [5′-TAGCGAGAGTTACTGTAACCA-3′] and AP1 adapter primer)–500 μM dNTP–50 mM Tris-HCl (pH 9.2)–14 mM (NH4 )SO4 –3 mM MgCl2 –2% dimethyl sulfoxide–0.1% Tween 20–2.5 U of Expand enzyme mixture (Boehringer Mannheim). .. The transcription plasmid used for in vitro translation of the wild-type H430 protein was constructed by insertion of an Apa I- Hin dIII fragment containing the entire H430 coding sequences into the Not I- Hin dIII site of the pTL2(glo) vector ( ).

Article Title: Globicatella bacteraemia identified by 16S ribosomal RNA gene sequencing
Article Snippet: The bacterial DNA extract and control were amplified with 0.5 μmol/l primers (LPW57 5′‐AGTTTGATCCTGGCTCAG‐3′ and LPW205 5′‐CTTGTTACGACTTCACCC‐3′; Gibco BRL, Rockville, MD, USA). .. The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mmol/l Tris‐HCl pH 8.3, 50 mmol/l KCl, 2 mmol/l MgCl2 , and 0.01% gelatin), 200 μmol/l of each dNTP and 1.0 U Taq polymerase (Boehringer Mannheim, Germany).

Article Title: Major Protein of Resting Rhizomes of Calystegia sepium (Hedge Bindweed) Closely Resembles Plant RNases But Has No Enzymatic Activity 1
Article Snippet: .. The reaction mixture for amplification of genomic DNA sequences contained 10 m m Tris-HCl, pH 8.3, 50 m m KCl, 1.5 m m MgCl2 , 100 mg/L gelatin, 0.4 m m of each dNTP, 2.5 units of Taq polymerase (Boehringer Mannheim, Basel), 50 ng to 500 μg of genomic DNA, and 20 μL of the appropriate primer mixture (20 μ m ) in a 100-μL reaction volume. ..

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: The bacterial DNA extracts and the control were amplified with 0.5 μM primers (LPW55 5′-AGTTTGATCCTGGCTCAG-3′ and LPW205 5′-CTTGTTACGACTTCACCC-3′) (Gibco BRL, Rockville, Md.). .. The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany).

Synthesized:

Article Title: Identification of a distinct class of cytoskeleton-associated mRNAs using microarray technology
Article Snippet: .. In addition, the abundance of histone 33, a transcript that distributed equally between the cytosolic and cytoskeleton fractions in microarray analysis, was confirmed. cDNA was synthesized from each sample by incubating 1 ug RNA sample with 0.5 mM each dNTP, 0.25 μg/ml random hexamer (Boehringer Mannheim), 10 U RNase inhibitor, and 10 U OmniScript reverse transcriptase in OmniScript RT Buffer (Qiagen). .. Two microliters of serially diluted template cDNA (1:3, 1:9, 1:27, 1:81, 1:243, 1:729 in ddH2 O) was used for each PCR reaction in a total volume of 24 μl containing 10 pmol of each primer, 0.2 mM each dNTP and 2.5 U Taq DNA polymerase in Qiagen PCR buffer.

Lambda DNA Preparation:

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany). .. Ten microliters of each amplified product was electrophoresed in 1.0% (wt/vol) agarose gel, with a molecular size marker (Lambda DNA Ava II digest; Boehringer Mannheim, Germany) in parallel.

Microarray:

Article Title: Identification of a distinct class of cytoskeleton-associated mRNAs using microarray technology
Article Snippet: .. In addition, the abundance of histone 33, a transcript that distributed equally between the cytosolic and cytoskeleton fractions in microarray analysis, was confirmed. cDNA was synthesized from each sample by incubating 1 ug RNA sample with 0.5 mM each dNTP, 0.25 μg/ml random hexamer (Boehringer Mannheim), 10 U RNase inhibitor, and 10 U OmniScript reverse transcriptase in OmniScript RT Buffer (Qiagen). .. Two microliters of serially diluted template cDNA (1:3, 1:9, 1:27, 1:81, 1:243, 1:729 in ddH2 O) was used for each PCR reaction in a total volume of 24 μl containing 10 pmol of each primer, 0.2 mM each dNTP and 2.5 U Taq DNA polymerase in Qiagen PCR buffer.

Incubation:

Article Title: Immunological and pathological comparative analysis between experimental latent tuberculous infection and progressive pulmonary tuberculosis
Article Snippet: Then, proteins were depleted by incubation with proteinase K 1 μg/ml (G ibco BRL) for 30 min at 37°C. .. The PCR was carried out by incubating the sections with 50 μl of 1× reaction buffer (G ibco BRL), 1·5 U Taq polymerase, 2 m m MgCl2 , 40 μ m dNTP, 0·2 μ m dUTP labelled with digoxigenin (Boehringer Manheim Indianapolis, IN, USA) and 60 pg each of IS6110 m tuberculosis insertion sequence primers.

Modification:

Article Title: Telomerase in kinetoplastid parasitic protozoa
Article Snippet: .. Ten microliters of the telomerase reaction was added to a 50-μl final volume PCR mix containing 1× modified TRAP buffer, 50 μM each dNTP, 20 pmol of TS primer, 20 pmol of CX-ext primer, 0.1 μCi/μl [α-32 P]dGTP or [α-32 P]dCTP and 1 unit of Taq polymerase (Boehringer Mannheim). .. PCR conditions and nuclease controls were the same as for the one-tube TRAP described above ( ).

Derivative Assay:

Article Title: Major Protein of Resting Rhizomes of Calystegia sepium (Hedge Bindweed) Closely Resembles Plant RNases But Has No Enzymatic Activity 1
Article Snippet: The reaction mixture for amplification of genomic DNA sequences contained 10 m m Tris-HCl, pH 8.3, 50 m m KCl, 1.5 m m MgCl2 , 100 mg/L gelatin, 0.4 m m of each dNTP, 2.5 units of Taq polymerase (Boehringer Mannheim, Basel), 50 ng to 500 μg of genomic DNA, and 20 μL of the appropriate primer mixture (20 μ m ) in a 100-μL reaction volume. .. The PCR primers were derived from both ends of the coding sequence of the cDNA clones encoding CalsepRRP.

Countercurrent Chromatography:

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: A GC-clamp (5′-CGC CCG CCG CGC CCC GCG CCC GTC CCG CCG CCC CCG CCC G-3′), which was added to the 5′ end of one primer in each pair ( GC ), made the resulting PCR products suitable for DGGE analysis (Myers et al. ; Sheffield et al. ; Abrams et al. ). .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.)

Immunohistochemistry:

Article Title: Immunological and pathological comparative analysis between experimental latent tuberculous infection and progressive pulmonary tuberculosis
Article Snippet: The same paraffin blocks as those used for the histopathology and immunohistochemistry were used for detection of mycobacterial DNA by in situ PCR [ ]. .. The PCR was carried out by incubating the sections with 50 μl of 1× reaction buffer (G ibco BRL), 1·5 U Taq polymerase, 2 m m MgCl2 , 40 μ m dNTP, 0·2 μ m dUTP labelled with digoxigenin (Boehringer Manheim Indianapolis, IN, USA) and 60 pg each of IS6110 m tuberculosis insertion sequence primers.

Telomerase Assay:

Article Title: Telomerase in kinetoplastid parasitic protozoa
Article Snippet: Ten microliters of the telomerase reaction was added to a 50-μl final volume PCR mix containing 1× modified TRAP buffer, 50 μM each dNTP, 20 pmol of TS primer, 20 pmol of CX-ext primer, 0.1 μCi/μl [α-32 P]dGTP or [α-32 P]dCTP and 1 unit of Taq polymerase (Boehringer Mannheim). .. To analyze the telomerase assay step separately from the PCR step, we also performed template-directed termination reactions in which the DEAE fractions were assayed under conditions in which the dNTPs were replaced by equimolar concentration of corresponding ddNTPs.

Generated:

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: These primer pairs were designed on the basis of the melt profiles generated by MacMelt Software (DNA Melt Profile Macintosh Software for the D Gene System, version 1.0) for the HV1 region (Steighner et al. ). .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.)

Polymerase Chain Reaction:

Article Title: Induction of neuropilin-1 and vascular endothelial growth factor by epidermal growth factor in human gastric cancer cells
Article Snippet: For reverse transcriptase–polymerase chain reaction (RT–PCR), 3 μ g of total RNA was used for cDNA synthesis with avian myeloblastosis virus reverse transcriptase (Life Technologies) in a final volume of 20 μ l. The reaction mixture included 0.5 M Tris-HCl (pH 8.0), 0.5 M KCl, 0.05 M MgCl2 , 2.5 mM dNTP, 40 U of RNase inhibitor (Boehringer Mannheim, Indianapolis, IN, USA), 50 U of reverse transcriptase, and 0.5 μ g of random primers. .. A portion of the reaction mixture (5 μ l) was subjected to PCR amplification in a reaction mixture (50 μ l) that contained 1 μ mol l−1 of each of two primers (sense and antisense), 1.5 mmol l−1 of MgCl2 , 0.2 mmol l−1 of each of four deoxynucleotides, and 2.5 U of Taq polymerase (Promega, Madison, WI, USA).

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.) .. PCR cycling consisted of initial denaturation at 95°C for 30 s, followed by 35 cycles consisting of denaturation at 95°C for 30 s, primer annealing at 50°C ( / GC ) or 60°C ( GC / ) for 30 s, and extension at 72°C for 30 s. Prior to DGGE analysis, the PCR products were heated to 95°C to denature the DNA and then were slowly cooled to room temperature to allow for reannealing of the DNA strands.

Article Title: Telomerase in kinetoplastid parasitic protozoa
Article Snippet: .. Ten microliters of the telomerase reaction was added to a 50-μl final volume PCR mix containing 1× modified TRAP buffer, 50 μM each dNTP, 20 pmol of TS primer, 20 pmol of CX-ext primer, 0.1 μCi/μl [α-32 P]dGTP or [α-32 P]dCTP and 1 unit of Taq polymerase (Boehringer Mannheim). .. PCR conditions and nuclease controls were the same as for the one-tube TRAP described above ( ).

Article Title: Serum levels of the angiogenic factor pleiotrophin in relation to disease stage in lung cancer patients
Article Snippet: .. PCR The cDNA (1.5 μl) was amplified in a 30 μl final volume of 10 mM Tris (pH 8.3), 50 mM KCl, 1.5 mM MgCl2 , 200 μl of each dNTP, 0.5 μM of each primer and 1.25 U Taq polymerase (Boehringer Mannheim, Germany) for 25–30 cycles under the following conditions: 5 min at 95°C, 1 min at 56.5°C (PTN) or at 58°C (GAPDH) and 2 min at 73°C. ..

Article Title: Polymorphisms of Human Leukocyte Antigen Genes in Korean Children with Kawasaki Disease
Article Snippet: .. The PCR procedure was carried out in a reaction (13 μl), containing 100 to 200 ng genomic DNA, 0.8 X buffer (40 mmol/l KCl, 1.2 mmol/l MgCl2 , 8 mmol/l Tris-HCl pH 8.8, 0.08% Triton X-100), 5% dimethylsulphoxide (DMSO), 200 μmol/l of each dNTP, 0.25 U Taq DNA polymerase (Boehringer, Mannheim, Germany), 1 μmol/l of each sequence-specific primer, and 0.2 μmol/l of internal control primers. .. The amplifications were performed in a My Cycler thermocycler (Bio-Rad, Hercules, USA).

Article Title: Immunological and pathological comparative analysis between experimental latent tuberculous infection and progressive pulmonary tuberculosis
Article Snippet: .. The PCR was carried out by incubating the sections with 50 μl of 1× reaction buffer (G ibco BRL), 1·5 U Taq polymerase, 2 m m MgCl2 , 40 μ m dNTP, 0·2 μ m dUTP labelled with digoxigenin (Boehringer Manheim Indianapolis, IN, USA) and 60 pg each of IS6110 m tuberculosis insertion sequence primers. .. The sequence of the primers was: 5-CCT GCG AGC GTA GGC GTC GG3-sense and 5-CTC GTC CAG CGC CGC TTC GG 3-antisense.

Article Title: Globicatella bacteraemia identified by 16S ribosomal RNA gene sequencing
Article Snippet: .. The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mmol/l Tris‐HCl pH 8.3, 50 mmol/l KCl, 2 mmol/l MgCl2 , and 0.01% gelatin), 200 μmol/l of each dNTP and 1.0 U Taq polymerase (Boehringer Mannheim, Germany). .. The mixtures were amplified in in an automated thermal cycler (Perkin‐Elmer Cetus, Gouda, The Netherlands), using 40 cycles of 94°C for 1 minute, 60°C for 1 minute, and 72°C for 2 minutes, with a final extension at 72°C for 10 minutes.

Article Title: Identification of a distinct class of cytoskeleton-associated mRNAs using microarray technology
Article Snippet: In addition, the abundance of histone 33, a transcript that distributed equally between the cytosolic and cytoskeleton fractions in microarray analysis, was confirmed. cDNA was synthesized from each sample by incubating 1 ug RNA sample with 0.5 mM each dNTP, 0.25 μg/ml random hexamer (Boehringer Mannheim), 10 U RNase inhibitor, and 10 U OmniScript reverse transcriptase in OmniScript RT Buffer (Qiagen). .. Two microliters of serially diluted template cDNA (1:3, 1:9, 1:27, 1:81, 1:243, 1:729 in ddH2 O) was used for each PCR reaction in a total volume of 24 μl containing 10 pmol of each primer, 0.2 mM each dNTP and 2.5 U Taq DNA polymerase in Qiagen PCR buffer.

Article Title: Major Protein of Resting Rhizomes of Calystegia sepium (Hedge Bindweed) Closely Resembles Plant RNases But Has No Enzymatic Activity 1
Article Snippet: Paragraph title: PCR Amplification of Genomic DNA Fragments Encoding CalsepRRP ... The reaction mixture for amplification of genomic DNA sequences contained 10 m m Tris-HCl, pH 8.3, 50 m m KCl, 1.5 m m MgCl2 , 100 mg/L gelatin, 0.4 m m of each dNTP, 2.5 units of Taq polymerase (Boehringer Mannheim, Basel), 50 ng to 500 μg of genomic DNA, and 20 μL of the appropriate primer mixture (20 μ m ) in a 100-μL reaction volume.

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: .. The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany). .. The mixtures were amplified in 40 cycles of 94°C for 1 min, 55°C for 1 min, and 72°C for 2 min, with a final extension at 72°C for 10 min in an automated thermal cycler (Perkin-Elmer Cetus, Gouda, The Netherlands).

DNA Sequencing:

Article Title: Globicatella bacteraemia identified by 16S ribosomal RNA gene sequencing
Article Snippet: PCR amplification and DNA sequencing of the 16S rRNA gene was performed according to our previous publications., Briefly, DNase I treated distilled water and PCR master mix (dNTPs, PCR buffer, and Taq polymerase) were used in all PCR reactions by adding 1 U of DNase I (Pharmacia, Sweden) to 40 μl of distilled water or PCR master mix, incubating the mixture at 25°C for 15 minutes, and subsequently at 95°C for 10 minutes to inactivate the DNase I. .. The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mmol/l Tris‐HCl pH 8.3, 50 mmol/l KCl, 2 mmol/l MgCl2 , and 0.01% gelatin), 200 μmol/l of each dNTP and 1.0 U Taq polymerase (Boehringer Mannheim, Germany).

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: PCR amplification and DNA sequencing of the 16S rRNA genes were performed as described in previous publications by members of our group ( , ). .. The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany).

Reverse Transcription Polymerase Chain Reaction:

Article Title: Induction of neuropilin-1 and vascular endothelial growth factor by epidermal growth factor in human gastric cancer cells
Article Snippet: .. For reverse transcriptase–polymerase chain reaction (RT–PCR), 3 μ g of total RNA was used for cDNA synthesis with avian myeloblastosis virus reverse transcriptase (Life Technologies) in a final volume of 20 μ l. The reaction mixture included 0.5 M Tris-HCl (pH 8.0), 0.5 M KCl, 0.05 M MgCl2 , 2.5 mM dNTP, 40 U of RNase inhibitor (Boehringer Mannheim, Indianapolis, IN, USA), 50 U of reverse transcriptase, and 0.5 μ g of random primers. ..

Article Title: Identification of a distinct class of cytoskeleton-associated mRNAs using microarray technology
Article Snippet: Paragraph title: RT-PCR ... In addition, the abundance of histone 33, a transcript that distributed equally between the cytosolic and cytoskeleton fractions in microarray analysis, was confirmed. cDNA was synthesized from each sample by incubating 1 ug RNA sample with 0.5 mM each dNTP, 0.25 μg/ml random hexamer (Boehringer Mannheim), 10 U RNase inhibitor, and 10 U OmniScript reverse transcriptase in OmniScript RT Buffer (Qiagen).

Antiviral Assay:

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany). .. Ten microliters of each amplified product was electrophoresed in 1.0% (wt/vol) agarose gel, with a molecular size marker (Lambda DNA Ava II digest; Boehringer Mannheim, Germany) in parallel.

Denaturing Gradient Gel Electrophoresis:

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: A GC-clamp (5′-CGC CCG CCG CGC CCC GCG CCC GTC CCG CCG CCC CCG CCC G-3′), which was added to the 5′ end of one primer in each pair ( GC ), made the resulting PCR products suitable for DGGE analysis (Myers et al. ; Sheffield et al. ; Abrams et al. ). .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.)

Cellular Antioxidant Activity Assay:

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: HV1 was analyzed as two overlapping fragments, by use of PCR primer pairs (5′-CCC AAA GCT AAG ATT CTA AT-3′)/ GC (5′- GC TGG CTT TGG AGT TGC AGT TG-3′) and GC (5′- GC TGA CCA CCT GTA GTA CAT AA-3′)/ (5′-GAG GAT GGT GGT CAA GGG AC-3′) (numbering system of Anderson et al. [ ]). .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.)

Article Title: Serum levels of the angiogenic factor pleiotrophin in relation to disease stage in lung cancer patients
Article Snippet: PCR The cDNA (1.5 μl) was amplified in a 30 μl final volume of 10 mM Tris (pH 8.3), 50 mM KCl, 1.5 mM MgCl2 , 200 μl of each dNTP, 0.5 μM of each primer and 1.25 U Taq polymerase (Boehringer Mannheim, Germany) for 25–30 cycles under the following conditions: 5 min at 95°C, 1 min at 56.5°C (PTN) or at 58°C (GAPDH) and 2 min at 73°C. .. Primer sequences were as follows: PTN, sense 5′ GGT CTC GAG TAT GTT CCA CAG GTG ACA TC 3′ and anti-sense 5′ GGT AAG CTT AGA GGA CGT TTC CAA CTC AA 3′; GAPDH sense 5′CGT CTT CAC CAT GGA GA 3′ and anti-sense 5′ GCC GGT AGT GCG GTG TCA AA 3′.

Histopathology:

Article Title: Immunological and pathological comparative analysis between experimental latent tuberculous infection and progressive pulmonary tuberculosis
Article Snippet: The same paraffin blocks as those used for the histopathology and immunohistochemistry were used for detection of mycobacterial DNA by in situ PCR [ ]. .. The PCR was carried out by incubating the sections with 50 μl of 1× reaction buffer (G ibco BRL), 1·5 U Taq polymerase, 2 m m MgCl2 , 40 μ m dNTP, 0·2 μ m dUTP labelled with digoxigenin (Boehringer Manheim Indianapolis, IN, USA) and 60 pg each of IS6110 m tuberculosis insertion sequence primers.

Nucleic Acid Electrophoresis:

Article Title: Globicatella bacteraemia identified by 16S ribosomal RNA gene sequencing
Article Snippet: Paragraph title: PCR, gel electrophoresis, and 16S ribosomal RNA gene sequencing ... The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mmol/l Tris‐HCl pH 8.3, 50 mmol/l KCl, 2 mmol/l MgCl2 , and 0.01% gelatin), 200 μmol/l of each dNTP and 1.0 U Taq polymerase (Boehringer Mannheim, Germany).

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: Paragraph title: PCR, gel electrophoresis, and 16S rRNA gene sequencing. ... The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany).

Mutagenesis:

Article Title: Polymorphisms of Human Leukocyte Antigen Genes in Korean Children with Kawasaki Disease
Article Snippet: HLA-A, -B, and -C Genotyping The genotyping of HLA-A, -B, and -C was performed by the amplification refractory mutation system (ARMS)–polymerase chain reaction (PCR) method. .. The PCR procedure was carried out in a reaction (13 μl), containing 100 to 200 ng genomic DNA, 0.8 X buffer (40 mmol/l KCl, 1.2 mmol/l MgCl2 , 8 mmol/l Tris-HCl pH 8.8, 0.08% Triton X-100), 5% dimethylsulphoxide (DMSO), 200 μmol/l of each dNTP, 0.25 U Taq DNA polymerase (Boehringer, Mannheim, Germany), 1 μmol/l of each sequence-specific primer, and 0.2 μmol/l of internal control primers.

Isolation:

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: Genomic DNA was isolated using either the Puregene DNA Isolation Kit (for whole blood; Gentra Systems), according to the manufacturer's protocol, or Chelex 100 chelating resin (for blood spots; Biorad) (Walsh et al. ; Holland et al. ). .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.)

Size-exclusion Chromatography:

Article Title: Identification of a distinct class of cytoskeleton-associated mRNAs using microarray technology
Article Snippet: In addition, the abundance of histone 33, a transcript that distributed equally between the cytosolic and cytoskeleton fractions in microarray analysis, was confirmed. cDNA was synthesized from each sample by incubating 1 ug RNA sample with 0.5 mM each dNTP, 0.25 μg/ml random hexamer (Boehringer Mannheim), 10 U RNase inhibitor, and 10 U OmniScript reverse transcriptase in OmniScript RT Buffer (Qiagen). .. PCR cycling conditions were 4 min at 94°C and then 24, 26 or 30 cycles of 45 sec at 94°C, 45 sec at 55–63°C and 1 min at 72°C.

Mouse Assay:

Article Title: Immunological and pathological comparative analysis between experimental latent tuberculous infection and progressive pulmonary tuberculosis
Article Snippet: Paragraph title: Mycobacterial DNA detection by in situ PCR in lungs from latent-infected mice ... The PCR was carried out by incubating the sections with 50 μl of 1× reaction buffer (G ibco BRL), 1·5 U Taq polymerase, 2 m m MgCl2 , 40 μ m dNTP, 0·2 μ m dUTP labelled with digoxigenin (Boehringer Manheim Indianapolis, IN, USA) and 60 pg each of IS6110 m tuberculosis insertion sequence primers.

Sequencing:

Article Title: Polymorphisms of Human Leukocyte Antigen Genes in Korean Children with Kawasaki Disease
Article Snippet: .. The PCR procedure was carried out in a reaction (13 μl), containing 100 to 200 ng genomic DNA, 0.8 X buffer (40 mmol/l KCl, 1.2 mmol/l MgCl2 , 8 mmol/l Tris-HCl pH 8.8, 0.08% Triton X-100), 5% dimethylsulphoxide (DMSO), 200 μmol/l of each dNTP, 0.25 U Taq DNA polymerase (Boehringer, Mannheim, Germany), 1 μmol/l of each sequence-specific primer, and 0.2 μmol/l of internal control primers. .. The amplifications were performed in a My Cycler thermocycler (Bio-Rad, Hercules, USA).

Article Title: Immunological and pathological comparative analysis between experimental latent tuberculous infection and progressive pulmonary tuberculosis
Article Snippet: .. The PCR was carried out by incubating the sections with 50 μl of 1× reaction buffer (G ibco BRL), 1·5 U Taq polymerase, 2 m m MgCl2 , 40 μ m dNTP, 0·2 μ m dUTP labelled with digoxigenin (Boehringer Manheim Indianapolis, IN, USA) and 60 pg each of IS6110 m tuberculosis insertion sequence primers. .. The sequence of the primers was: 5-CCT GCG AGC GTA GGC GTC GG3-sense and 5-CTC GTC CAG CGC CGC TTC GG 3-antisense.

Article Title: Globicatella bacteraemia identified by 16S ribosomal RNA gene sequencing
Article Snippet: Paragraph title: PCR, gel electrophoresis, and 16S ribosomal RNA gene sequencing ... The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mmol/l Tris‐HCl pH 8.3, 50 mmol/l KCl, 2 mmol/l MgCl2 , and 0.01% gelatin), 200 μmol/l of each dNTP and 1.0 U Taq polymerase (Boehringer Mannheim, Germany).

Article Title: Major Protein of Resting Rhizomes of Calystegia sepium (Hedge Bindweed) Closely Resembles Plant RNases But Has No Enzymatic Activity 1
Article Snippet: The reaction mixture for amplification of genomic DNA sequences contained 10 m m Tris-HCl, pH 8.3, 50 m m KCl, 1.5 m m MgCl2 , 100 mg/L gelatin, 0.4 m m of each dNTP, 2.5 units of Taq polymerase (Boehringer Mannheim, Basel), 50 ng to 500 μg of genomic DNA, and 20 μL of the appropriate primer mixture (20 μ m ) in a 100-μL reaction volume. .. The PCR primers were derived from both ends of the coding sequence of the cDNA clones encoding CalsepRRP.

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: Paragraph title: PCR, gel electrophoresis, and 16S rRNA gene sequencing. ... The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany).

Chloramphenicol Acetyltransferase Assay:

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: HV1 was analyzed as two overlapping fragments, by use of PCR primer pairs (5′-CCC AAA GCT AAG ATT CTA AT-3′)/ GC (5′- GC TGG CTT TGG AGT TGC AGT TG-3′) and GC (5′- GC TGA CCA CCT GTA GTA CAT AA-3′)/ (5′-GAG GAT GGT GGT CAA GGG AC-3′) (numbering system of Anderson et al. [ ]). .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.)

Article Title: Serum levels of the angiogenic factor pleiotrophin in relation to disease stage in lung cancer patients
Article Snippet: PCR The cDNA (1.5 μl) was amplified in a 30 μl final volume of 10 mM Tris (pH 8.3), 50 mM KCl, 1.5 mM MgCl2 , 200 μl of each dNTP, 0.5 μM of each primer and 1.25 U Taq polymerase (Boehringer Mannheim, Germany) for 25–30 cycles under the following conditions: 5 min at 95°C, 1 min at 56.5°C (PTN) or at 58°C (GAPDH) and 2 min at 73°C. .. Primer sequences were as follows: PTN, sense 5′ GGT CTC GAG TAT GTT CCA CAG GTG ACA TC 3′ and anti-sense 5′ GGT AAG CTT AGA GGA CGT TTC CAA CTC AA 3′; GAPDH sense 5′CGT CTT CAC CAT GGA GA 3′ and anti-sense 5′ GCC GGT AGT GCG GTG TCA AA 3′.

Rapid Amplification of cDNA Ends:

Article Title: Characterization of SRp46, a Novel Human SR Splicing Factor Encoded by a PR264/SC35 Retropseudogene
Article Snippet: Rapid amplification of cDNA ends (RACE)-PCR experiments were performed with human normal thymus polyadenylated mRNA species and the Marathon cDNA amplification kit (Clontech) according to the manufacturer’s instructions. .. Double-stranded cDNA was amplified in the presence of a 0.8 μM concentration of each amplimer (H430-1507 [5′-TAGCGAGAGTTACTGTAACCA-3′] and AP1 adapter primer)–500 μM dNTP–50 mM Tris-HCl (pH 9.2)–14 mM (NH4 )SO4 –3 mM MgCl2 –2% dimethyl sulfoxide–0.1% Tween 20–2.5 U of Expand enzyme mixture (Boehringer Mannheim).

Software:

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: These primer pairs were designed on the basis of the melt profiles generated by MacMelt Software (DNA Melt Profile Macintosh Software for the D Gene System, version 1.0) for the HV1 region (Steighner et al. ). .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.)

Negative Control:

Article Title: Globicatella bacteraemia identified by 16S ribosomal RNA gene sequencing
Article Snippet: The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mmol/l Tris‐HCl pH 8.3, 50 mmol/l KCl, 2 mmol/l MgCl2 , and 0.01% gelatin), 200 μmol/l of each dNTP and 1.0 U Taq polymerase (Boehringer Mannheim, Germany). .. DNase I treated distilled water was used as the negative control.

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany). .. DNase I-treated distilled water was used as the negative control.

Agarose Gel Electrophoresis:

Article Title: Serum levels of the angiogenic factor pleiotrophin in relation to disease stage in lung cancer patients
Article Snippet: PCR The cDNA (1.5 μl) was amplified in a 30 μl final volume of 10 mM Tris (pH 8.3), 50 mM KCl, 1.5 mM MgCl2 , 200 μl of each dNTP, 0.5 μM of each primer and 1.25 U Taq polymerase (Boehringer Mannheim, Germany) for 25–30 cycles under the following conditions: 5 min at 95°C, 1 min at 56.5°C (PTN) or at 58°C (GAPDH) and 2 min at 73°C. .. PCR products were loaded on a 1.5% ethidium bromide-stained agarose gel.

Article Title: Polymorphisms of Human Leukocyte Antigen Genes in Korean Children with Kawasaki Disease
Article Snippet: The PCR procedure was carried out in a reaction (13 μl), containing 100 to 200 ng genomic DNA, 0.8 X buffer (40 mmol/l KCl, 1.2 mmol/l MgCl2 , 8 mmol/l Tris-HCl pH 8.8, 0.08% Triton X-100), 5% dimethylsulphoxide (DMSO), 200 μmol/l of each dNTP, 0.25 U Taq DNA polymerase (Boehringer, Mannheim, Germany), 1 μmol/l of each sequence-specific primer, and 0.2 μmol/l of internal control primers. .. The presence or absence of PCR products was determined after separation of the samples on a 1.5% agarose gel containing 0.5 μg/ml of ethidium bromide.

Article Title: Globicatella bacteraemia identified by 16S ribosomal RNA gene sequencing
Article Snippet: The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mmol/l Tris‐HCl pH 8.3, 50 mmol/l KCl, 2 mmol/l MgCl2 , and 0.01% gelatin), 200 μmol/l of each dNTP and 1.0 U Taq polymerase (Boehringer Mannheim, Germany). .. A 10 μl sample of each amplified product was electrophoresed in 1.5% (w/v) agarose gel, with a molecular size marker (φX174 Hae III digest,; Boehringer Mannheim, Germany) in parallel.

Article Title: Identification of a distinct class of cytoskeleton-associated mRNAs using microarray technology
Article Snippet: In addition, the abundance of histone 33, a transcript that distributed equally between the cytosolic and cytoskeleton fractions in microarray analysis, was confirmed. cDNA was synthesized from each sample by incubating 1 ug RNA sample with 0.5 mM each dNTP, 0.25 μg/ml random hexamer (Boehringer Mannheim), 10 U RNase inhibitor, and 10 U OmniScript reverse transcriptase in OmniScript RT Buffer (Qiagen). .. PCR products were separated by agarose gel electrophoresis, stained with ethidium bromide and visualized in a FluorImager (Molecular Dynamics, Sunnydale, CA).

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany). .. Ten microliters of each amplified product was electrophoresed in 1.0% (wt/vol) agarose gel, with a molecular size marker (Lambda DNA Ava II digest; Boehringer Mannheim, Germany) in parallel.

In Situ:

Article Title: Immunological and pathological comparative analysis between experimental latent tuberculous infection and progressive pulmonary tuberculosis
Article Snippet: Paragraph title: Mycobacterial DNA detection by in situ PCR in lungs from latent-infected mice ... The PCR was carried out by incubating the sections with 50 μl of 1× reaction buffer (G ibco BRL), 1·5 U Taq polymerase, 2 m m MgCl2 , 40 μ m dNTP, 0·2 μ m dUTP labelled with digoxigenin (Boehringer Manheim Indianapolis, IN, USA) and 60 pg each of IS6110 m tuberculosis insertion sequence primers.

Random Hexamer Labeling:

Article Title: Identification of a distinct class of cytoskeleton-associated mRNAs using microarray technology
Article Snippet: .. In addition, the abundance of histone 33, a transcript that distributed equally between the cytosolic and cytoskeleton fractions in microarray analysis, was confirmed. cDNA was synthesized from each sample by incubating 1 ug RNA sample with 0.5 mM each dNTP, 0.25 μg/ml random hexamer (Boehringer Mannheim), 10 U RNase inhibitor, and 10 U OmniScript reverse transcriptase in OmniScript RT Buffer (Qiagen). .. Two microliters of serially diluted template cDNA (1:3, 1:9, 1:27, 1:81, 1:243, 1:729 in ddH2 O) was used for each PCR reaction in a total volume of 24 μl containing 10 pmol of each primer, 0.2 mM each dNTP and 2.5 U Taq DNA polymerase in Qiagen PCR buffer.

DNA Extraction:

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: Genomic DNA was isolated using either the Puregene DNA Isolation Kit (for whole blood; Gentra Systems), according to the manufacturer's protocol, or Chelex 100 chelating resin (for blood spots; Biorad) (Walsh et al. ; Holland et al. ). .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.)

Concentration Assay:

Article Title: A Sensitive Denaturing Gradient-Gel Electrophoresis Assay Reveals a High Frequency of Heteroplasmy in Hypervariable Region 1 of the Human mtDNA Control Region
Article Snippet: .. PCR amplification was performed by combining 10–50 ng DNA template with 1 × Pfu reaction buffer (20 mM Tris-HCl pH 8.8, 2 mM MgSO4 , 10 mM KCl, 10 mM (NH4 )2 SO4 , 0.1% Triton X-100, 0.1 mg nuclease-free BSA per ml), 200 μM each dNTP (Boehringer Mannheim), 0.2 μM each primer (Synthetic Genetics), 2 μg BSA (Sigma Chemical), 2.5 U Pfu polymerase, and sufficient sterile distilled water (Gibco BRL Life Technologies) to bring the final volume to 50 μl. (The accepted SI unit of concentration, mol/L, has been represented by the symbol “M,” in order to conform to the conventions of this journal.) .. PCR cycling consisted of initial denaturation at 95°C for 30 s, followed by 35 cycles consisting of denaturation at 95°C for 30 s, primer annealing at 50°C ( / GC ) or 60°C ( GC / ) for 30 s, and extension at 72°C for 30 s. Prior to DGGE analysis, the PCR products were heated to 95°C to denature the DNA and then were slowly cooled to room temperature to allow for reannealing of the DNA strands.

Article Title: Telomerase in kinetoplastid parasitic protozoa
Article Snippet: Ten microliters of the telomerase reaction was added to a 50-μl final volume PCR mix containing 1× modified TRAP buffer, 50 μM each dNTP, 20 pmol of TS primer, 20 pmol of CX-ext primer, 0.1 μCi/μl [α-32 P]dGTP or [α-32 P]dCTP and 1 unit of Taq polymerase (Boehringer Mannheim). .. To analyze the telomerase assay step separately from the PCR step, we also performed template-directed termination reactions in which the DEAE fractions were assayed under conditions in which the dNTPs were replaced by equimolar concentration of corresponding ddNTPs.

Article Title: Characterization of SRp46, a Novel Human SR Splicing Factor Encoded by a PR264/SC35 Retropseudogene
Article Snippet: .. Double-stranded cDNA was amplified in the presence of a 0.8 μM concentration of each amplimer (H430-1507 [5′-TAGCGAGAGTTACTGTAACCA-3′] and AP1 adapter primer)–500 μM dNTP–50 mM Tris-HCl (pH 9.2)–14 mM (NH4 )SO4 –3 mM MgCl2 –2% dimethyl sulfoxide–0.1% Tween 20–2.5 U of Expand enzyme mixture (Boehringer Mannheim). .. The transcription plasmid used for in vitro translation of the wild-type H430 protein was constructed by insertion of an Apa I- Hin dIII fragment containing the entire H430 coding sequences into the Not I- Hin dIII site of the pTL2(glo) vector ( ).

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: .. The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany). .. The mixtures were amplified in 40 cycles of 94°C for 1 min, 55°C for 1 min, and 72°C for 2 min, with a final extension at 72°C for 10 min in an automated thermal cycler (Perkin-Elmer Cetus, Gouda, The Netherlands).

Marker:

Article Title: Globicatella bacteraemia identified by 16S ribosomal RNA gene sequencing
Article Snippet: The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mmol/l Tris‐HCl pH 8.3, 50 mmol/l KCl, 2 mmol/l MgCl2 , and 0.01% gelatin), 200 μmol/l of each dNTP and 1.0 U Taq polymerase (Boehringer Mannheim, Germany). .. A 10 μl sample of each amplified product was electrophoresed in 1.5% (w/v) agarose gel, with a molecular size marker (φX174 Hae III digest,; Boehringer Mannheim, Germany) in parallel.

Article Title: Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis
Article Snippet: The PCR mixture (50 μl) contained bacterial DNA, PCR buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2 mM MgCl2 , and 0.01% gelatin), a 200 μM concentration of each dNTP, and 1.0 U of Taq polymerase (Boehringer Mannheim, Mannheim, Germany). .. Ten microliters of each amplified product was electrophoresed in 1.0% (wt/vol) agarose gel, with a molecular size marker (Lambda DNA Ava II digest; Boehringer Mannheim, Germany) in parallel.

Staining:

Article Title: Identification of a distinct class of cytoskeleton-associated mRNAs using microarray technology
Article Snippet: In addition, the abundance of histone 33, a transcript that distributed equally between the cytosolic and cytoskeleton fractions in microarray analysis, was confirmed. cDNA was synthesized from each sample by incubating 1 ug RNA sample with 0.5 mM each dNTP, 0.25 μg/ml random hexamer (Boehringer Mannheim), 10 U RNase inhibitor, and 10 U OmniScript reverse transcriptase in OmniScript RT Buffer (Qiagen). .. PCR products were separated by agarose gel electrophoresis, stained with ethidium bromide and visualized in a FluorImager (Molecular Dynamics, Sunnydale, CA).

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    Boehringer Mannheim dntp
    Dntp, supplied by Boehringer Mannheim, used in various techniques. Bioz Stars score: 92/100, based on 95 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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