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Chapter 3 Mate-Paired Library Preparation
Overview
3
Overview
For 2 × 50 bp mate-paired libraries, size-selected genomic DNA fragments are ligated
to LMP CAP Adaptors and circularized with internal adaptors (see Figure 7). The
resulting DNA circle has one nick in each strand because the LMP CAP Adaptor does
not have the 5′ phosphate in one of its oligonucleotides. Nick translation using E. coli
DNA polymerase I “pushes” the nick into the genomic DNA region in 5′ to 3′
direction. The length of nick-translated DNA can be controlled by adjusting reaction
temperature and time. T7 exonuclease and S1 nuclease digestion cuts the DNA at the
position opposite to the nick and releases the DNA mate pair. P1 and P2 Adaptors are
then ligated to the ends of the mate-paired library for subsequent amplification by PCR
(see Figure 9 on page 45).
Figure 7
Basic 2 × 50 bp mate-paired library preparation workflow.
SOLiD™ 4 System Library Preparation Guide
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Chapter 3 Mate-Paired Library
Preparation
This chapter describes the method to make a mate-paired library with insert sizes
ranging from 600 bp to 6 kb. A mate-paired library consists of pairs of DNA fragments
that are “mates” because they originated from the two ends of the same genomic DNA
fragment. CAP adaptors connect the DNA mate pair together through an internal
adaptor.