User guide

59
Fragment Library Preparation Using the AB Library Builder
™
System: 5500 Series SOLiD
™
Systems User Guide
C
APPENDIXC
Supplemental Background
Information
This appendix covers:
■ Why prepare fragment libraries? . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
■ Preparing fragment libraries. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 59
■ Sequence orientation from source DNA to sequence map. . . . . . . . . . . . . . . . . . . 62
Why prepare fragment libraries?
Features
• Appropriate for sequence lengths ≤ 300 bp.
• Adaptors on each end of sheared DNA insert.
• Multiplexed sequencing.
• The protocol is designed for 10 ng–5 µg of genomic DNA.
• Compared to mate-paired libraries, fragment libraries yield a higher recovery of
unique molecules, when normalized to the same input amount.
Applications
• Targeted resequencing, primary library
• Genomic resequencing
• Methylation analysis
Complexity
The amount of library used depends on the application and information needed. For
deeper coverage of large and complex genomes (for example, human genomes), more
DNA is required to prepare libraries. For smaller and less complex genomes (for
example, microbial genomes), less DNA can be used. For information about specific
applications, go to the 5500 Series SOLiD
™
Sequencers website:
www.appliedbiosystems.com/solid5500
Or, contact your field applications specialist.
Preparing fragment libraries
Fragment library preparation involves shearing DNA into small fragments and
ligating P1-T and barcoded adaptors specific for fragment library preparation (see
Figure 1 on page 60).