Human parvovirus B19 NS1 gene for non-structural protein NS1, partial cds, isolate: N81.
Submission Date
1999-07-29
Strain Name
N81
Sampling Date
1983
Sampling Country
Japan
Location
Japan
Submitted By
Ishii,K.K.
Submitting Institution
Contact:Keiko K Ishii Tohoku University School of Medicine, Department of Molecular Diagnostics; 1-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8574, Japan
Used in the mixture for incubating sera to extract B19 DNA.
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10 mM-Tris-HCl (pH 7.5)
Buffer component used during the extraction of B19 DNA.
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1 mM-EDTA
Component in the enzyme mixture for DNA extraction.
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0.5% SDS
Detergent used in the mixture for DNA extraction.
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Phenol, Phenol-chloroform (1:1), and Chloroform
Used in sequential extraction steps to purify B19 DNA.
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Ethanol
For precipitating DNA during extraction.
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Distilled water
Used to dissolve the precipitated DNA for PCR.
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PCR primers (various)
Specifically, primers Prm 4-a, Prm 4-b, Prm 6-a, Prm 8-b, Prm 8-a, Prm 10-b, Prm 10-a, and Prm 6-b are mentioned for amplifying different regions of the B19 genome.
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dATP, dCTP, dGTP, dTTP (100 μM each)
Nucleotide building blocks for DNA amplification in PCR.
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10 mM-Tri~HCl (pH 9.0)
Buffer component for PCR reaction.
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2 mM-MgCl2
Essential cofactor for the Taq DNA polymerase enzyme.
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50 mM-KCl
Component in the PCR buffer to maintain ionic strength.
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0.1% Triton X-100
Non-ionic surfactant used in PCR reactions.
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Taq DNA polymerase (2 units, Promega)
Enzyme used to synthesize new DNA strands in PCR.
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Mineral oil
Overlay to prevent evaporation during PCR.
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PstI and DraI restriction enzymes
Used to cleave PCR amplified B19 DNA fragments at specific sites.
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5% acrylamide gel
Medium for separating DNA fragments during gel electrophoresis.
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M13mpl0 or M13mpll phage vector
Vectors used for cloning the PCR-amplified B19 DNA fragments.
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Hybrid plasmids (pUK372, pUK370, pUK371)
Vectors constructed for cloning the various PCR products encompassing almost the entire B19 genome.
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SmaI site of plasmid pUC18
Site used for cloning PCR products for further analysis.
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Klenow fragment
Used to prepare PCR products before cloning into plasmids.
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Set of restriction endonucleases recognizing six base pair sequences
Used for analysis of restriction site polymorphisms (including BamHI, BglII, BstEII, EcoRI, HindIII, HpaI, KpnI, PstI, PvuII, SalI, SmaI, XbaI, XhoI).
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Set of restriction endonucleases recognizing 4- or 5-base pair sequences
Used for more detailed genomic analysis (including AluI, Cfr13I, DdeI, HaeIII, HhaI, HinclI, HinfI, MboII, MspI, RsaI, Sau3AI, TaqI).
