User guide

16
Fragment Library Preparation Using the AB Library Builder
™
System: 5500 Series SOLiD
™
Systems User Guide
Chapter2 Prepare to Build the Library
Shear the DNA
b. Set the chiller temperature to 2–5°C to ensure that the temperature reading in
the water bath displays 5°C.
c. Supplement the circulated water chiller with 20% ethylene glycol.
2. Dilute the desired amount of DNA to 120 µL in 1✕ Low TE Buffer in a LoBind
tube. Shear Buffer reduces DNA damage from fragmentation:
3. Load the DNA into the Covaris
®
S2 System:
a. Place a Covaris
®
microTUBE into the loading station.
b. Keeping the snap-cap on the tube, use a tapered pipette tip to slowly transfer
the 120 µL of DNA sample through the pre-split septa.
Be careful not to introduce a bubble into the bottom of the tube.
Note: To load and unload the Covaris
®
microTUBE correctly from the
microTUBE holder, see “Load and unload Covaris
®
microTUBE vials from
the Covaris
®
microTUBE holder” on page 47.
4. Shear the DNA using the following Covaris
®
S2 System conditions:
IMPORTANT! Ensure that the bath temperature limit is set at 15°C, and keep the
bath temperature to ≤ 10°C.
5. Remove the sheared DNA:
a. Place the Covaris
®
microTUBE into the loading station.
b. While keeping the snap-cap on, insert a pipette tip through the pre-split
septa, then slowly remove the sheared DNA.
c. Transfer 110 µL of the sheared DNA into a new 1.5-mL sample tube
provided in the Library Builder
™
Fragment Core Kit for SOLiD
™
4.0.
Component Amount
DNA 10 ng to 5 µg
1✕ Low TE Buffer Variable µL
Shear Buffer 1.2 µL
Total 120 µL
Condition Setting
Number of cycles 6
Bath temperature 5°C
Bath temperature limit 15°C
Mode Frequency
sweeping
Water quality testing function Off
Duty cycle 10%
Intensity 5
Cycles/burst 100
Time 60 seconds