Polymerase Chain Reaction:Article Title: Methylation of INK4 and CIP/KIP families of cyclin-dependent kinase inhibitor in chronic lymphocytic leukaemia in Chinese patients
Article Snippet: .. The polymerase chain reaction (PCR) mixture contained 50 ng of bisulphite‐treated DNA, 0.2 mM deoxynucleoside triphosphates, 2 mM magnesium chloride, 10 pmol of each primer, 1 × PCR buffer II and 2.5 units of AmpliTaq Gold (Perkin‐Elmer Biosystems, Wellesley, Massachusetts, USA) in a final volume of 50 μl. ..
Article Title: Fas/Fas ligand (FasL)-deregulated apoptosis and IL-6 insensitivity in highly malignant myeloma cells
Article Snippet: .. 935) were the internal primers. cDNA (100 ng) and 50 pmol of each primer were added to the polymerase chain reaction (PCR) mixture (Perkin Elmer Cetus, Norwalk, CT) with subsequent amplification in a thermal cycler for 35 cycles (1 min 94°C/1 min 58°C/1 min 72°C), which in our system were found to provide suitable amounts of PCR product. .. This was then analysed on a 2% agarose gel, cloned in the pCRTM II vector (Invitrogen) and amplified in DH5α bacterial strain (G ibco BRL, Milan, Italy) for sequencing by high-voltage electrophoresis on a 6% urea-bis-acrylamide gel with the Sequenase 2.0 kit (Amersham, Aylesbury, UK).
Article Title: Mitochondrial decay in hepatocytes from old rats: Membrane potential declines, heterogeneity and oxidants increase
Article Snippet: .. The polymerase chain reaction (PCR) mixture consisted of 10 mM Tris·HCl (pH 8.3), 50 mM KCl, 0.001% gelatin, 200 μM each dNTP, 2.5 mM MgCl2 , 1.25 units of Ampli Taq DNA polymerase (Perkin–Elmer), 10 pmol of each primer, and 500 ng of total DNA in 50 μl; for 1 cycle of 94°C for 2 min, followed by 40 cycles of 94°C for 60 sec, 60°C for 30 sec, and 72°C for 60 sec, and 1 cycle of 72°C for 5 min. Primer 1 (5′-GCTTAGAGCGTTAACCTTTTAAG-3′; nucleotides 7687–7709) and primer 2 (5′-CAGCAGTTTGGATGTGGCG-3′; nucleotides 12962–12981) were used for detection of the 16-nt direct repeat-associated mtDNA deletion at nucleotides 8103 and 12937 ( ) of the rat mtDNA sequence. .. Primer 3 (5′-GGTTCTTACTTCAGGGGCCATC-3′; nucleotides 15768–15789) and primer 4 (5′-GTGGAATTTTCTGAGGGTAGGC-3′; nucleotides 16268–16288) were used for the amplification of wild-type mtDNA.
Sequencing:Article Title: Mitochondrial decay in hepatocytes from old rats: Membrane potential declines, heterogeneity and oxidants increase
Article Snippet: .. The polymerase chain reaction (PCR) mixture consisted of 10 mM Tris·HCl (pH 8.3), 50 mM KCl, 0.001% gelatin, 200 μM each dNTP, 2.5 mM MgCl2 , 1.25 units of Ampli Taq DNA polymerase (Perkin–Elmer), 10 pmol of each primer, and 500 ng of total DNA in 50 μl; for 1 cycle of 94°C for 2 min, followed by 40 cycles of 94°C for 60 sec, 60°C for 30 sec, and 72°C for 60 sec, and 1 cycle of 72°C for 5 min. Primer 1 (5′-GCTTAGAGCGTTAACCTTTTAAG-3′; nucleotides 7687–7709) and primer 2 (5′-CAGCAGTTTGGATGTGGCG-3′; nucleotides 12962–12981) were used for detection of the 16-nt direct repeat-associated mtDNA deletion at nucleotides 8103 and 12937 ( ) of the rat mtDNA sequence. .. Primer 3 (5′-GGTTCTTACTTCAGGGGCCATC-3′; nucleotides 15768–15789) and primer 4 (5′-GTGGAATTTTCTGAGGGTAGGC-3′; nucleotides 16268–16288) were used for the amplification of wild-type mtDNA.
Amplification:Article Title: Fas/Fas ligand (FasL)-deregulated apoptosis and IL-6 insensitivity in highly malignant myeloma cells
Article Snippet: .. 935) were the internal primers. cDNA (100 ng) and 50 pmol of each primer were added to the polymerase chain reaction (PCR) mixture (Perkin Elmer Cetus, Norwalk, CT) with subsequent amplification in a thermal cycler for 35 cycles (1 min 94°C/1 min 58°C/1 min 72°C), which in our system were found to provide suitable amounts of PCR product. .. This was then analysed on a 2% agarose gel, cloned in the pCRTM II vector (Invitrogen) and amplified in DH5α bacterial strain (G ibco BRL, Milan, Italy) for sequencing by high-voltage electrophoresis on a 6% urea-bis-acrylamide gel with the Sequenase 2.0 kit (Amersham, Aylesbury, UK).
Size-exclusion Chromatography:Article Title: Mitochondrial decay in hepatocytes from old rats: Membrane potential declines, heterogeneity and oxidants increase
Article Snippet: .. The polymerase chain reaction (PCR) mixture consisted of 10 mM Tris·HCl (pH 8.3), 50 mM KCl, 0.001% gelatin, 200 μM each dNTP, 2.5 mM MgCl2 , 1.25 units of Ampli Taq DNA polymerase (Perkin–Elmer), 10 pmol of each primer, and 500 ng of total DNA in 50 μl; for 1 cycle of 94°C for 2 min, followed by 40 cycles of 94°C for 60 sec, 60°C for 30 sec, and 72°C for 60 sec, and 1 cycle of 72°C for 5 min. Primer 1 (5′-GCTTAGAGCGTTAACCTTTTAAG-3′; nucleotides 7687–7709) and primer 2 (5′-CAGCAGTTTGGATGTGGCG-3′; nucleotides 12962–12981) were used for detection of the 16-nt direct repeat-associated mtDNA deletion at nucleotides 8103 and 12937 ( ) of the rat mtDNA sequence. .. Primer 3 (5′-GGTTCTTACTTCAGGGGCCATC-3′; nucleotides 15768–15789) and primer 4 (5′-GTGGAATTTTCTGAGGGTAGGC-3′; nucleotides 16268–16288) were used for the amplification of wild-type mtDNA.
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