Product Manual
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10
7.4 BUFFER PREPARATION AND PARAMETERS
For all protocols prepare the following:
Lysis Buffer PA2
Check for precipitated SDS especially if stored below 20°C. If necessary, incubate the bottle
for several minutes at 30‑40°C and thoroughly mix until SDS precipitate is re‑dissolved
completely.
Preparing Wash Buffer PAW2 with ethanol
Add 96‑100% ethanol to Wash Buffer PAW2 Concentrate: 24 mL for the 10 prep kit, 100 mL
for the 50 prep kit and 200 mL for the 250 prep kit.
Important Note:
Mark bottle label to indicate ethanol was added. Wash Buffer PAW2 at is stable at room
temperature (18‑25°C) for at least one year
Preparing RNase A stock solution
Reconstitute lyophilized RNase A in nuclease‑free water: 150 µL for the 10 prep kit, 0.6 mL
for the 50 prep kit and 1.5 mL x 2 for the 250 prep kit. Store RNase A solution at 4°C for up to
3 months. For long‑term storage of up to 1 year, divide into small aliquots and store at ‑20°C.
Elution Parameters
Elute DNA using Elution Buffer PG (supplied). In general, two consecutive elutions increases
DNA yield compared to a single elution at the same total buffer volume. This is particularly
important for small buffer volumes. However, large volumes of elution buffer or the use of two
consecutive elutions will result in low DNA concentration. The standard elution procedure
is already optimized to yield 80–90% after two elution steps at elevated temperatures. The
elution protocol can be modified to improve yield, concentration or increase elution speed:
•
Standard elution: dispense 50 μL Elution Buffer PG, incubate for 5 min at 65°C and
repeat (85‑90% yield).
•
Maximum yield: dispense 100 μL Elution Buffer PG, incubate for 5 min at 65°C and
repeat (95‑100% yield).
•
High concentration: dispense 25 μL Elution Buffer PG, incubate for 5 min at 65°C and
repeat (75% yield).
• Rapid elution: dispense 100 μL Elution Buffer PG, incubate for 1‑5 min at room
temperature or 65°C (60‑70% yield).
Summary of Contents for ISOLATE II Plant DNA Kit
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Page 17: ...Plant DNA Kit 17 NOTES...
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