User guide
22
Fragment Library Preparation Using the AB Library Builder
™
System: 5500 Series SOLiD
™
Systems User Guide
Chapter2 Prepare to Build the Library
Set up the AB Library Builder
™
System for size-selected or express fragment library preparation
b. Calculate the amount of each P1-T and barcoded adaptor (Y) needed. Y=µL
of P1-T=µL of barcoded adaptor:
where:
c. Calculate the amount of 1✕ Low TE Buffer required, based on the volumes of
P1-T and barcoded adaptors needed
d. Add 1✕ Low TE Buffer, P1 and barcoded adaptors to the appropriate 5✕
Reaction Buffer tube. Vortex, then pulse-spin the tube and place it in open
position 11 of the cartridge.
Load Agencourt AMPure
®
XP Reagent in the cartridges
Thoroughly resuspend Agencourt AMPure
®
XP Reagent, then carefully transfer
500 µL to unsealed position 12.
Load the tip and tube rack
IMPORTANT! If you are processing fewer than 13 samples, make sure to load the tips
and tubes in the same positions as the reagent cartridges that are loaded in the
cartridge rack.
Load the tip and tube rack in the following order:
1. Row S (fourth row): Load with sample/elution tubes containing 110 µL of
sheared DNA. If the sample volume is <110 µL, then add 1✕ TE to the sample for
a total volume of 110 µL. Ensure that the cap is off.
2. Row T1 and T2 (second and third rows): Load with AB Library Builder
™
Tips
inserted into tip holders.
Note: Two tip and tip holder sets are required per sample.
A = (value below), if... Library type
0.3 Fragment
0.66 Express
fragment
Y µL adaptor = (value below), if...
0.56 µL (1:10 dilution) <100 ng
(fragment
library)
1.21 µL (1:10 dilution) <100 ng
(express
fragment
library
Y μL adaptor needed = # μg DNA ×
9.2 pmol
1 μg DNA
× 10 ×
1 μL adaptor needed
50 pmol
A ×
μL 1X Low TE Buffer needed =
42.5 – 2Y