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1
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Histopaque 1119 (Sigma Diagnostics)
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To isolate leukocytes from the bone marrow sample.
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10405389
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2
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150 mM NaCl
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To wash the leukocytes after isolation.
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3
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Buffer (10 mM Tris-HCl [pH 8.3], 50 mM KCl, 2.5 mM MgCl2, 0.5% Tween 20, 0.5% Nonidet P-40)
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To lyse the leukocyte cells.
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4
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Proteinase K (15 mg)
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To digest proteins during the cell lysis.
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5
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Phenol
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To extract proteins from the lysate.
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6
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Chloroform-isoamyl alcohol (25/1)
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To further remove proteins from the lysate.
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7
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Glycogen (20 mg as the carrier)
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To aid in the precipitation of DNA.
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8
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3 M Sodium acetate (pH 5.2)
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To adjust the ionic strength for DNA precipitation.
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9
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Ethanol
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To precipitate DNA from the solution.
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10
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70% Ethanol
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To wash the DNA pellet.
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11
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H2O
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To resuspend the DNA pellet.
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12
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QIAamp Blood Kit (Qiagen)
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To extract viral DNA from serum samples.
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13
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Restriction enzyme (BamHI, HindIII, or PvuII)
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To digest double-stranded DNA for restriction mapping.
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14
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Formamide
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Component of the hybridization buffer.
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15
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53 SSC (0.75 M NaCl plus 0.075 M sodium citrate)
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Component of the hybridization buffer.
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16
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Blocking reagent (Boehringer)
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Component of the hybridization buffer to prevent non-specific binding.
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17
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N-laurylsarcosine
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Component of the hybridization buffer.
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18
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Sodium dodecyl sulfate (SDS)
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Component of the hybridization buffer.
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19
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B19-specific probe, P2560 labeled with the Dig Oligonucleotide Tailing Kit (Boehringer)
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To hybridize with the B19 PCR products.
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20
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Alkaline phosphatase conjugate
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For detection as part of the DIG Luminescent Detection Kit (Boehringer).
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21
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CSPD (DIG Luminescent Detection Kit; Boehringer)
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For chemiluminescent detection of the hybridized probe.
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22
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Taq DyeDeoxy Terminator Cycle Sequencing Kit (Perkin-Elmer Applied Biosystems)
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For sequencing PCR products.
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